Biochem Image Bank Edition 2 Updated Upto NEET PG 21

Biochemistry
Image Bank
Index
Sl.No. Chapter Pg.No.
1. Sub cellular organelles and Cell membrane 09
2. Amino Acids 10
3. Proteins 23
4. Enzymes 36
5. Carbohydrates 42
6. Absorption of Carbohydrates 51
7. Metabolism of Carbohydrates 52
8. Lipids 68
9. Cholesterol and Lipoproteins 77
10. Lipid Storage Disorders 82
11. Molecular Genetics - Structure of DNA 85
12. DNA Replication 87
13. Different Classes of RNA 92
14. Epigenetics 93
15. Recombinant DNA Technology 94
16. Amplification and Hybridisation Techniques 98
17. Pattern of Inheritance 101
18. Immunochemistry 102
19. Vitamins and Minerals 105
20. Heme Synthesis and Breakdown 114
21. Electron Transport Chain 118

9
Sub cellular organelles

and
Cell membranes
Sub cellular organelles and marker enzymes
Sub cellular Marker Enzyme
organelle
Mitochondria Inner membrane;
ATP synthase
Lysosome Cathepsin
Golgi complex Galactosyl transferase
Microsomes Glucose-6-transferase
Cytoplasm Lactate dehydrogenase
Reference : Textbook of Biochemistry for Medical Students by DM Vasudevan 7th Ed. Pg 11
Fluid mosaic model

•
Lipid bilayer originally proposed by Davson and
Danielle was later described as a fluid mosaic
model by Singer and Nicolson.
•
The distribution of the phospholipids is such that
choline containing phospholipids are mainly in the
external layer and ethanolamine and serine
containing phospholipids in the inner layer.
•
Each leaflet is 25 A thick, with the head portion
10 A and tail 15 A thick. The total thickness is
about 50 to 80 A.
•
This flip-flop movement is catalyzed by enzymes.
•
Flippases catalyze the transfer of amino phospholipids across the membrane.
•
Floppases catalyze the outward directed movement, which is ATP dependent.
•
The nature of the fattyacids also affects the fluidity of the membrane, the more unsaturated cis fatty
acids increase the fluidity. Reference : Textbook of Biochemistry for Medical Students by DM Vasudevan 7th Ed. Pg 14
10
Amino Acids
Chemistry of amino acids
Spectrophotometry
•
Quantitative estimation of Proteins
•
Proteins will absorb ultraviolet light at (250 - 290nm).
(Amino acids donot absorb visible light)
•
This is due to the tryptophan(major contribution),tyrosine
and phenylalanine residues in the protein.
@
Tryptophan absorbs in region of 280nm. NEET ‘20
coauthor
Titration curve
Titration curves are useful in identification of Amino acids.
Reference : Harper’s Illustrated Biochemistry 31st Ed. Pg20.
@
Amino acids exists as Ampholytes or Zwitter ions in solution depending on pH of solution.
@
At Isoelectric pH (pI) - No net charge,Solubitily and buffering capacity minimum.
@
In Acidic medium they exist in Cationic.

@
In Basic medium they exist in Anionic.
At points pK they resist change in pH in addition of an acid or base (Buffering capacity).
:From the graph of Glycine,its evident that there are 2 pK values

one at lower pH ( Amino grp )and one in higher pH ( Carboxyl grp ),which means 2 ionisable group.
So maximum buffering capacity will be around these 2 points (i.e pK1 and pK2) for Glycine(contains
only 2 ionisable grp )
o
Histidine has an extra ionisable grp other than amino and carboxyl grp ,hence 3 pK values.
Titration curve Histidine Titration curve Glycine AIIMS ‘19

11
structure of amino acid
General structure
Most of the amino acids are Alpha
0
COO - Carboxyl group
amino acids where Amino group and
NH3 - Amino group
:R - R group or
Carboxyl group are attached to the
same C atom except Proline which
Side chain Imino acid.
Simple amino acid
Glycine ( Gly) Alanine ( Ala)
Branched amino acids
Valine ( Val ) Leucine ( Leu ) Isoleucine ( Ile)

12
Hydroxyamino acids Sulfur containing amino acids
Serine ( Ser) Threonine ( Thr ) Cysteine ( Cys ) Methionine ( Met )
Amino acids with amino group Dicarboxylic amino acids
Glutamine ( Gln ) Aspartic acid ( Asp ) Glutamic Acid ( Glu)

Asparagine ( Asn )
Dibasic amino acids Aromatic amino acids
Phenylalanine ( Phe )
Arginine ( Arg ) contains Lysine ( Lys )

Guanidium group Tyrosine ( Tyr )
13
Amino acid with special group
Tryptophan ( Trp ) Histidine ( His )
Indole group Imidazole group

Proline ( Pro )
O
The Alpha amino nitrogen
form a rigid five membered
Pyrrolidine ring.
some Derived amino acids
4- Hydroxy Proline
Gamma carboxy Ornithine

Glutamic acid
14
GENERAL AMINO ACID METABOLISM
UREA CYCLE DISORDERS
Carbamoyl phosphate Ornithine

Transcarbamoylase
Citrulline
Ornithine
Ornithine Transcarbamoylase deficiency
@
Only urea cycle disorder with X Linked inheritance
@
Hyperammonemia Type II NEET ‘19
@
Elevated Ammonia with High Glutamine level in Blood and CSF
Trichorrhexis nodosa Nitrogen Scavengers used in Urea Cycle Disorder

o
Phenylbutyrate form Phenylbutylglutamine. AIIMS ‘19 ‘17
s/i Arginosuccinate lyase deficiency o
Benzoate forms Hippuric acid.
o
Phenylacetate form Phenylacetylglutamine.
15
AROMATIC AMINO ACIDS
Pheochromocytoma
A Catecholamine producing tumor of Sympathetic and Parasympathetic
o
Nervous System.
Catecholamines are degraded by 2 enzymes COMT and MAO,
o
NEET ‘ 18
Vanillyl Mandelic Acid ( VMA ) is produced in excess which is excreted in urine.
“ Rule of Ten “ - 10% B/L , 10% Extra-adrenal , 10% Malignant
o
Clinical feature
Palpitations
Ams
Headaches
Ams
Profuse Sweating
Ams
Investigations
24 hr Urine VMA
o
Metanephrines ( Total and Fractionated )

o
Plasma metanephrines
o
PHENYLKETONURIA AIIMS ‘18

•
Caused by Phenylalanine hydroxylase deficiency

•
Recessive condition
Clinical Manifestation
Mental Retardation
•
Mousy Body Odour due to
•
Agitation
•
Phenyllactic acid
Hyperactivity
•
Tremors
•
Hypopigmented hair Convulsions

•
Hypopigmentation
•
Diagnosis
Blood phenylalanine - level >20mg/dl
•
Tandem Mass Spectrometry

•
Ferric Chloride test

•
DNA Probes
•
1 - Phenylalanine hydroxylase
1A - NADPH dependent reductase
Treatment Reference : Textbook of
Diet containing low Phenylalanine

•
Guthrie Test Biochemistry for Medical Students
by DM Vasudevan 7th Ed. Pg 236
16
ALKAPTONURIA
o
Autosomal Recessive. NEET ‘20
o
Caused by Homogentisic acid oxidase deficiency.
o
Blackening of urine on standing
o
Ochronosis - accumulation of black polymer of
homogentisic acid.
o
Arthritis - Acute exacerbations present.
Radiologic features similar to Osteoarthritis.
Diagnosis
Massive excretion of homogentisic acid on urine testing.
o
Tyrosine levels - Normal.

Dark spots on sclera and cartilage
o
Benedict’s test - Strongly Positive.

o
Ferric Chloride test positive.

o
Treatment
Symptomatic
o
Nitisinone combined with Phenylalanine and Tyrosine

o
Blackening of Urine restricted diet.

Nelson Testbook of Pediatrics 20th Ed. Pg 642
A patient complains of knee pain. Routine investigations are unremarkable and still, the patient is unsatisfied. Urine
turns black on standing, what is the enzyme involved ?
A. Homogentisate oxidase C. Tyrosine transaminase
B. Xanthine oxidase D. 4- fumarylacetoacetase
Albinism
NEET ‘19
Autosomal Recessive Disease
: Absent Tyrosinase - defective synthesis of Melanin.
Clinical Manifestations
i. Hypopigmented ocular fundus
ii. Red or Gray iris
iii. Photophobia
iv. Nystagmus
v. Decreased visual acquity
vi. Nevi and Melanomas in skin
Red Iris
Reference : Nelson Textbook of Pediatrics 30th Ed Pg 642
17
Classification of Albinism
Oculocutaneous Albinism OCULAR Albinism

i.OCA 1 - Tyrosinase Negative •
Nettleship - Falls Type
ii.OCA 2 - Tyrosinase Positive
iii.OCA 3 - Rufos Albinism
iv.OCA 4
Syndromes associated
Most common type among Generalised OCA is : OCA 2 i. Hermansky Pudlak Syndrome
ii. Chediak - Higashi syndrome
Localised albinism
@
Localised patches of hypopigmentation of skin and hair
Piebaldism
Autosomal Dominant
: Born with white forelock
White macules on the face,trunk and extremities.
: Mutation in KIT gene present.
Waardenburg syndrome
Autosomal Dominant
: Four Major Types :
Type I - Most common
: Type II
Type III
Type IV - Associated with Hirschsprung disease
White forelock
i
Lateral displacement of inner canthi of eyes
Broad Nasal Bridge
Heterochromia of Irides
Sensorineural deafness
@
18
Cystinuria
•
Autosomal Recessive
•
Inborn error in metabolism included in Garrod’s Tetrad
•
Epithelial cells of urine prevents absorption of
cystine,arginine,lysine,ornithine.( due to deficiency in
transport system.)
•
Cystine along with these amino acids may be excreted in urine.
Garrod’s Tetrad
Cystine crystals in Urine
i. Albinism iii.Cystinuria
ii.Alkaptonuria iv.Pentosuria.
Investigation
Cyanide Nitroprusside test - Postive
•
Urine - Acidic
•
Sulphur content in Cystine makes crystals radio-opaque.

•
Treatment
Supplement with Fluids
•
Cyanide Nitroprusside test showing Alkalanisation of urine by Sodium bicarbonate

•
Magenta Red Coloured Complex Reference : Textbook of Biochemistry for Medical Students by DM Vasudevan 7th Ed. Pg 219
Nelson Textbook of Pediatrics 20th Ed. Pg 2602
TRYPTOPHAN METABOLISM
NEET ‘ 18
Acetyl Serotonin
5 HT Serotonin
( 5-Hydroxytryptamine )
5 HIAA
19
Carcinoid Tumors
a.k.a Argentaffinomas
: Tumors develop in small intestine or appendix
CLINICAL MAnifestations
Flushing
:
Sweating
Intermittent diarrhoea
: Fluctuating hypertension
Picture of carcinoid ( at center) Diagnosis

encroaches into lumen of small bowel i. Serum Serotonin level increased
NEET ‘ 18
ii. 5 HIAA level in urine - increased
Hartnup disease
Autosomal Recessive
: Defect in the transport of neutral amino acids
including tryptophan by intestinal mucosa and
renal tubules.
Mostly asymptomatic
: Cutaneous photosensitivity
Pellagra like rash
: Pruritis
Chronic Eczema
Chronic eczema
: Intermittent ataxia
Diagnosis Irritability
i. Urinary aminoaciduria restricted to : Emotional instability
neutral amino acids Suicidal tendencies
ii.Plasma concentration of
Neutral Amino acids are usually normal :
Treatment
Nicotinic acid or nicotinamide
iii.Urinary excretion of proline,hydroxyproline High protein diet
0
and arginine normal

Reference : Nelson Testbook of Pediatrics 20th Ed. Pg 648
20
Blue diaper syndrome

a.k.a Indicanuria or Drummond syndrome
: Tryptophan is specifically malabsorbed (Defect in absorption only in intestine)
o
Unabsorbed tryptophan is converted to indican ( blue colour ) by gut bacteria.
o
Vomiting Failure to thrive
o
o
Constipation Nephrocalcinosis
o
8
Poor appetite irritability
•
classic Homocystinuria
Most common inborn error of methionine metabolism.
: Autosomal Recessive trait
Deficient Cystathionine Beta synthase ( Chromosome 21 )
: Clinical manifestation
Failure to thrive
Developmental delay
: Ectopic lentis ( Subluxation of Ocular lens )
Ectopia lentis Severe Myopia and iridodonesis
: Progressive intellectual disability - common
Psychiatric and behavioural disorders
: Convulsions
Skeletal deformities resembling Marfan syndrome
: Scoliosis,Pectus excavatum or carinatum
Arachnodactyly
: Thromboembolic episodes
Diagnosis
i. Methionine and Homocystine levels are
elevated in body fluids.( Diagnostic lab findings)
ii.Cystine level is low or absent in plasma
iii. Enzyme assay
Pectus Carinatum Treatment
11. In a patient with cystathionine B synthase i. Vitamin B6
deficiency, which aminoacid supplementation ii. Betaine - lowers homocysteine levels in
should be given? body fluids.
A. Methionine C. Cysteine Reference : Nelson Testbook of Pediatrics 20th Ed. Pg 644
B. Serine D. Tyrosine [NEET 2021]
21
Maple syrup urine disease
Autosomal Recessive
: Decarboxylation of leucine,isoleucine and
valine is done by BCKDH system.
Branched Chain alpha Ketoacid
: Dehydrogenase (TPP is co enzyme)
Deficiency any of the 4 subunits of
enzyme cause MSUD.
Type I - due to E1 alpha subunit
: Type II - due to E1 beta subunit
Type III - due to E2 subunit
Control Positive ( Yellow precipitate ) : Type IV - due to E3 subunit
2,4 Dinitrophenylhydazine test
clinical manifestations Diagnosis
o
Normal at birth Maple syrup odour of urine,sweat and cerumen
o
o
Poor feeding and Vomiting Urine - high levels of leucine,isoleucine and valine and
o
o
Lethargy and Coma respective ketoacids.
o
Severe Opisthotonus Positive 2,4-Dinitrophenylhydrazine test for ketoacids
o
Convulsions Neuroimaging - Cerebral edema

o
:
Treatment
In acute state - hydration and rapid removal of branched chain amino acids
Mannitol,diuretics or hypertonic saline used if cerebral edema is present.
:
Diet low in branched chain amino acids
Liver transplantation
Isovaleric acidemia
Autosomal Recessive,IVD gene on Chr 15
: Deficiency of Isovaleryl CoA dehydrogenase.
“Sweaty feet odour” in body sweat and cerumen not in urine.
: Vomiting,Severe acidosis
Lethargy, Convulsions, Coma
:
Diagnosis
Ketoacidosis 0
Elevated level of Isovaleric acid and metabolites in urine
Hypocalcemia Elevated level of Isovalerylcarnitine in plasma
: Hyperglycemia : Measurement of Enzyme activity in cultured skin fibroblast.
22
Canavan disease
o
Autosomal Recessive
o
Deficiency of enzyme Aspartoacylase
o
N-Aceytlaspartic acid is reservoir for acetate needed
for myelin synthesis.
o
Aspartoacylase cleaves N- Acetyl aspartic acid.
o
Spongy degenration of white matter
Appear normal at birth
o
Progressive macrocephaly
o
Severe hypotonia
Macrocephaly : Persistent Head lag
Delayed mile stones.
o
Optic atrophy
o
Seizures
o
Feeding difficulties
o
Diagnosis
o
CT and MRI shows diffuse white matter
degeneration primarily in cerebral
hemispheres with less involvement of
cerebellum and brainstem.
o
Elevated levels of N-acetyl aspartic acid in
the urine or blood ( Definitive diagnosis )
Treatment
No specific treatment is available
o
Genetic counselling,Carrier testing and

o
Axial MRI showing extensive Prenatal diagnosis - methods of prevention.

thickening of white matter.
23
Proteins
Structural organization of protein
Primary structure •
Denotes number and sequence of Amino
acids in proteins.
•
Higher levels organisation decided by
primary structure.
•
Each polypeptide has unique arrangement
of amino acids.
•
Maintained by Covalent bonds of peptide
linkage.
Secondary structure
Denotes the configuration also relationship
•
between residues which are about 3-4 Amino

acids apart in linear sequence.
Preserved by non covalent forces like
•
i. Hydrogen bond
ii. Ionic bonds
iii. Hydrophobic bonds
EERIE iv. Van der Walls forces
Alpha helix
Beta pleated sheet
Most common and stable form
•
Distance between adjacent AA residues is 3.5 A : Stabilized by Hydrogen bonds
Stabilized by Hydrogen bonds.
Each turn is formed by 3.6 residues.
•
Beta bends formed are stabilised by Intrachain

Distance between each AA residue is 1.5 A
•
disulfide bridges.
Proline and Hydroxyproline will not allow to
•
form Alpha helix.

Tertiary structure NEET ‘20
•
Denotes 3D structure of Protein
•
Defines steric relationship of AA which are far apart.
•
Maintained by Non - covalent forces.
•
Domain is the term used to denote a compact globular
functional unit of a protein.(independent region - may
represent functional unit)

24
Quarternary structure
•
Polypeptides aggregate to form one functional protein.
•
Loses its function when the subunits are dissociated.
•
Preserved by non covalent forces.
•
Depending upon polypeptide chain present : protein may
be termed as Monomer,Dimer,Tetramer etc.
•
Homodimer - 2 copies of same polypeptide chain.
•
Heterodimer - 2 different types of polypeptides.
Eg:
i. Creatine kinase - Dimer
ii. LDH - Tetramer
# Myoglobin - Alpha helix and Bet a pleated sheet.

# Collagen - Triple helix.
# Elastin - No specific structural motif.
TANDEM MASS SPECTROMETRY

•
Used to analyse complex peptide mixtures without

prior purification.
•
2 mass spectrometers linked in series.
•
Used to screen blood samples for abnormalities in
metabolites.
•
Applications :
i. Identification and quantification of
proteins.
MS - Mass Spectrometry ii. Drug Screening.
iii. Pesticides and pollutants AIIMS ‘18
iv. Screening of inborn errors of metabolism
Harper’s Illustrated Biochemistry 31st Ed. Pg 30
9.Which is the most accepted method for HbA1c estimation ?

A.Affinity chromatography
B.Ion exchange chromatography
C.Immunoturbimetry
D.Capillary electrophoresis [INICET 2021]
25
•
Determination of hemoglobin A1c (HbA1c) is one of the most important monitoring procedures for
long-term control of diabetes mellitus.
Several analytical methods have been developed for the measurement of glycohemoglobin (GHb).
: Those most frequently used are ion-exchange chromatography for HbA1c and affinity
chromatography for total GHb.
Biochemical techniques
Ion exchange chromatography AIIMS ‘18
•
Based on Electrostatic attraction between
-
A charged biological molecules to oppositely
charged groups on the ion exchange resins.
B
Resins are either Cation or Anion exchange resins.
→
i Separation is based on the ionic character of
proteins and amino acids ( iso-electric points).
•
Clinical assays are carried out by this method.
Ion exchange chromatography # Routine clinical assays are now carried out by
A - Negatively charged molecules attach with beads
High Performance Liquid Chromatography.
B - Positively charged molecules repel and
moves down faster Reference : Textbook of Biochemistry for Medical Students by DM Vasudevan 7th Ed.
Pg 329,451
Size-exclusion chromatography
Separates proteins according to size.
: The column matrix is a cross-linked polymer with

pores of selected size.
Larger proteins migrate faster than smaller ones,
because they are too large to enter the pores in
the beads and hence take a more direct route
through the column.
•
The smaller proteins enter the pores and are
slowed by their more labyrinthine path through
the column.
ooo
Reference : Lehninger Principles of Biochemistry 4th edition Pg. 91
26
Affinity chromatography
•
Separates proteins by their binding
specificities.
•
The proteins retained on the column are
those that bind specifically to a ligand
cross-linked to the beads.
•
After proteins that do not bind to the
ligand are washed through the column,
the bound protein of particular interest
is eluted (washed out of the column) by
a solution containing free ligand.
Electrophoresis
Different samples are loaded in wells or
i
depressions at the top of the polyacrylamide gel.
The proteins move into the gel when an electric
field is applied.
Proteins can be visualized after
electrophoresis by treating the gel with a
stain such as Coomassie blue, which binds to
the proteins butnot to the gel itself.
@
Each band on the gel represents a different
protein (or protein subunit); smaller proteins
move through the gel more rapidly than
larger proteins and therefore are found
nearer the bottom of the gel.
27
Isoelectric focusing
@
Is a procedure used to determine the Isoelectric
point (pI) of a protein.
A stable pH gradient is established in the gel
A
after application of an electric field.
Protein solution is added and electric
B
field is reapplied.
After staining proteins are shown to be

C distributed along pH gradient according to
A B C their pI values.
Fibrous proteins
Collagen
Collagen types seen
Type I - Most abundant,connective tissues.
•
Type II - Cartilage and vitreous humor.

•
Type III - Skin,lung and vascular tissues.

•
Type IV - Basement membrane.

•
Post-translational processing of Collagen
odessa
Edens
Abnormalities in Collagen cause AIIMS ‘18

i. Osteogenesis imperfecta •
Type I, III and V are True fibrillar
ii. Ehlers-Danlos Syndrome- Defective Type III collagen
collagen form main connective
iii. Alport syndrome - Abnormal Type IV collagen
tissue during healing wounds.
iv. Epidermolysis bullosa - Defective type VII collagen
Reference : Textbook of Biochemistry for Medical Students by DM Vasudevan 7th Ed. Pg 720,Harsh Mohan Textbook of Pathology 7th Ed. Pg.160
28
Fibrillin
Are Structural components of microfibrils.
:
Secreted into Extracellular matrix by fibroblast
Fibrillin microfibrils are found in elastic fibers and elastin free bundles in eye,kidney and tendons.
Marfan syndrome
Autosomal Dominant trait
: Caused by mutations in the gene ( on Chr 15 ) for Fibrillin - 1.
Ectopia lentis
: Arachnodactyly
Hyperextensibility of joints
:
Dilatations of Ascending Aorta
Tall patients
Hyperextensibility of joints
Arachnodactyly Ectopia lentis

Reference : Harper’s Illustrated Biochemistry 31st Edition Pg 597
29
Protein folding and degradation
Proteasome
NEET ‘20
•
Macromolecule, ubiquitous in nature.
•
Involved in Degradation of ubiquitin tagged protein occurs.
•
Proteins are degraded into small oligopeptides of 5-6

amino acids in length.
•
For degradation protein molecule must enter core,which is
regulated by 2 outer rings that recognise
polyubiquitinated proteins.
Reference : Textbook of Biochemistry for Medical Students by DM Vasudevan 7th Ed. Pg.199
Colour reactions of amino acids and proteins

Ninhydrin reaction
All amino acids when heated with ninhydrin can form complexes;
@
pink, purple or blue in color.

The color complex is called Ruhemann’s purple.
@
Proline and hydroxy proline will give yellow color with ninhydrin.
Amino acids with amide group (glutamine, asparagine) produce a
@
brown color.
Qualitative as well as quantitative estimation of amino acids.
@
Biuret test
@
Cupric ions in an alkaline medium form a violet color with

peptide bond nitrogen.
@
Individual amino acids and dipeptides will not answer this test.
Used for quantitative estimation also.
@
Magnesium and ammonium sulfates interfere with this reaction.

30
Xanthoproteic Test
@
The ring systems in phenylalanine, tyrosine and
tryptophan undergo nitration on treatment with
concentrated nitric acid when heated.
@
The end product is yellow in color which is intensified in
strong alkaline medium.
@
This reaction causes the yellow stain in skin by nitric
acid.
Negative Test Positive Test

Millon’s Test
@
Phenol group of phenylalanine and tyrosine containing proteins,
when heated with mercuric sulfate in sulfuric acid and sodium
nitrite (or, mercurous and mercuric nitrates in nitric acid) form
red colored mercury phenolate.
@
Chloride interferes with this reaction and so it is not suitable to
test for tyrosine in urine samples.
Sakaguchi’s Test Pauly’s Test

•
Diazo-benzene sulfonic
@
Free arginine or arginyl
acid reacts with imidazole
residues in proteins react
group of Histidine to form
with alpha-naphthol and
a cherry-red colored
alkaline hypobromite to
diazotized product under
give bright-red color.
alkaline conditions.
@
This is due to the •
Phenol group of tyrosine
guanidium group.
will give an orange red
colored product.

31
Sulfur Test for Cysteine
@
When cysteine or cysteine-containing proteins are boiled with
strong alkali, organic sulfur splits and forms sodium sulfide,
which on addition of lead acetate produces lead sulphide as a
black precipitate.
@
Methionine does not answer.
@
Albumin and keratin will answer sulfur test positive.
@
Casein will give a negative test.
Negative Positive Reference : Textbook of Biochemistry for Medical Students by DM Vasudevan 7th Ed. Pg 32
Nitroprusside Reaction
•
Proteins with free sulfhydryl groups give a reddish color
with sodium nitroprusside, in ammoniacal solution.
•
Many proteins give a negative, reaction in the native
state, but when denatured, reaction will be positive
Cyanide Nitroprusside test showing

Magenta Red Coloured Complex
32
Plasma proteins
Normal ELECPHOREtic pattern of plasma proteins
Separated bands of protein
Densitometer scanning reveals the relative mobilities of

albumin, α1-globulin, β2-globulin, β-globulin, and γ-globulin.
Normal ELECPHOREtic pattern of plasma proteins

Protein losing enteropathy
Loss of albumin and gamma globulins
att
: Slight increase in the Alpha 2 band is
due to a stressful stimulus
33
Nephrotic syndrome
Smaller proteins are lost more rapidly than large
molecular weight protein like Alpha 2
An
Macroglobulin.
There is decreased Albumin ,Alpha-1,Beta and
Gamma fraction.
Slight increase in Alpha 2 band because of
inflammation and stress
MOnoclonal Gammopathy
Multiple Myeloma
It
: Colonial synthesis of a unique
immunoglobulin,results in a sharp increase of
Gamma globulin peak ( called as M band ).
A-
Slight decrease in albumin.
Hepatic cirrhosis
Polyclonal gammopathy
Decreased Albumin ( due to decreased synthesis )
An
To balance this there is increase in
Gamma globulin ( to maintain Oncotic pressure )
Beta - Gamma bridging can be seen.
34
Immediate response pattern
•
Acute Phase Response
•
Stress or Inflammation caused by infection,injury or
Ham
surgical trauma cause change in the following proteins.
•
Acute Phase proteins that are increased are
i. CRP
ii. Haptoglobulin
iii. Macroglobulin
iv. Ceruloplasmin
Late response pattern

Chronic inflammation
:
Associated with infection shows an
t.no
increase in the gamma globulin peak
due to increase in immunoglobulins.
Alpha 2 band is also increased and
albumin peak is decreased.
Immunosuppressive diseases
There is decrease in gamma globulin peak.
A-
https://memorang.com/flashcards/108161/Blood,+Plasma+and+Serum+Components
35
Bence jones proteinuria
0
Seen in 20% of patients with multiple myeloma.
Monoclonal light chains are excreted in urine ( due
:
to asynchronous production of H and L chains or due
to deletion of portions of L chains, so that they
cannot combine with H chains )
Bence-Jones proteins have the special property of
precipitation when heated between 45 C and 60 C;
but redissolving at higher than 80 C and lower than
Crystal of Bence Jones protein 45 C.
These proteins may block kidney tubules, leading to
: renal failure.
Bradshaw’s test is also positive,when a few mL of
urine is layered over a few mL of concentrated
hydrochloric acid, a white ring of precipitate is
formed.
Waldenstorm’s macroglobulinemia
Ig M level in blood is increased with monoclonal peak ( due to malignant proliferation
: of Ig M clones).
Males are affected mostly.
Serum Viscosity is increased ( Ig M being macromolecules they form aggregates )
: SIA Test Positive
a
Recurrent bleeding is seen in patients due to hyperviscosity serum.
36
Enzymes
General enzymology
Gibbs free energy curve AIIMS ‘20
G= H- T S
•
G = Change in free energy
•
H = Change in Enthalpy
•
S = Change in Entropy
•
It denotes a portion of total energy change

in a system that is available for doing work.
•
The activation energies, G , for the S P and P S reactions are indicated.
G is the overall standard free-energy change in the direction S P.
10
Factors influencing enzyme activity

1. Enzyme concentration 6. Presence of activators
2. Substrate concentration 7. Presence of inhibitors
3. Product concentration 8. Presence of repressor or derepressor
4. Temperature 9. Covalent modification.
5. Hydrogen ion concentration (pH)
Enzyme concentration
•
Rate of a reaction or velocity (V) is directly
proportional to the enzyme concentration, when
sufficient substrate is present.
Reference : Textbook of Biochemistry for Medical Students by DM Vasudevan 7th Ed. Pg. 56
37
Substrate concentration
-
•
As substrate concentration is increased,
the velocity is also correspondingly
increased in the initial phases; but
the curve flattens afterwards.
•
Maximum velocity obtained is called Vmax
•
Represents the maximum reaction rate
attainable in presence of excess
substrate (at substrate saturation level).
michaelis constant
•
Km value is substrate concentration (expressed in moles/L) at half-maximal velocity.
•
It denotes that 50% of enzyme molecules are bound with substrate molecules at that particular
•
substrate concentration.
•
Km is independent of enzyme concentration.
•
Km is characteristic feature of a particular enzyme for a specific substrate.
•
The affinity of an enzyme towards its substrate is inversely related to the dissociation constant
Km denotes the affinity of enzyme for substrate. The lesser the numerical value of Km, the affinity of the
enzyme for the substrate is more.
CO-OPERATIVE BINDING
•
Not strictly follow the Michaelis-Menten kinetics.
•
binding of substrate to one unit enhances the affinity
for binding to other subunits.
•
Hill equation is employed.
•
Example :
i. Oxygen binding to Hemoglobin
38
Effect of temperature
•
Velocity of enzyme increases when
temperature increases reaches maximum
and then falls.
•
Temperature at which maximum amount of
the substrate is converted to the product
per unit time is called the Optimum
temperature.
Bell Shaped Curve
•
The temperature coefficient (Q10) is the
factor by which the rate of catalysis is
increased by a rise in 10°C.
Effect ph
•
Each enzyme has an optimum pH, on

both sides of which the velocity will be
drastically reduced.
•
Optimum pH may vary depending on
the temperature, concentration of
substrate, presence of ions, etc.
Bell Shaped Curve

Enzyme inhibition
Uncompetitive inhibition
•
Inhibitor binds to enzyme–substrate
complex; but not to the free enzyme.
•
Both Vmax and Km are Decreased
Eg :
Inhibition of placental alkaline
phosphatase (Regan iso-enzyme) by
phenylalanine.
Lineweaver-Burk plot of Uncompetitive inhibition.
39
Competitive inhibition
Lineweaver-Burk plot •
Inhibitor molecules are competing with the normal

substrate molecules for attaching with the active
site of the enzyme.
•
Effective concentration of enzyme is reduced, the
reaction velocity is decreased.
•
Inhibitor will be a structural analog of the
substrate.
•
Usually Reversible or increase in substrate
concentration abolishes inhibition.
•
Km value is Increased
•
Vmax value is Unchanged
Substrate saturation curve
Examples :
i. Sulfa drugs inhibit bacterial folate synthesis v. Allopurinol inhibits Xanthine oxidase used in Gout
ii. Methotrexate inhibit Folate reductase enzyme vi. Lovastatin inhibits HMGCoA reductase
iii. Dicoumarol inhibiting Vit K activity vii. Oseltamivir inhibits Neuraminidase
iv. Isoniazid structurally similar to Pyridoxal. viii. 5 FU inhibits Thymidilate synthase
Non competitive inhibition NEET ‘20

Lineweaver-Burk plot •
Inhibitor usually binds to a different domain on the
enzyme, other than the substrate binding site.
•
No structural resemblance to the substrate, an
increase in the substrate concentration generally does
not relieve this inhibition.
•
Km value Unchanged
•
Vmax value is Reduced
•
Examples :
i.Cyanide inhibits cytochrome oxidase.
ii.Fluoride inhibits Enolase
iii.Iodoacetate would inhibit enzymes having -SH
group in their active centers.
iv.BAL is used as antidote for heavy metal
Substrate saturation curve poisoning.
40
Allosteric regulation of enzymes
Allosteric enzyme
has one catalytic site where
•
the substrate binds and

another separate allosteric
site where the modifier binds.
Substrate saturation curve

Allosteric inhibition
Binding of Regulatory molecule with Allosteric site
•
inhibits binding of Substrate with Enzyme.

Examples :
•
i. ATP acts as an allosteric inhibitor (negative modifier)

of Phosphofructokinase 1.
ii. The end product, heme will allosterically inhibit the
ALA synthase.
iii. Cytidine triphosphate (CTP) the end product will
allosterically inhibit aspartate transcarbamoylase.
Allosteric activator NEET ‘ 18

•
Binding of Activator to Allosteric site enables binding of
substrate to active site of enzyme
•
Examples:
i. Aspartate transcarbamoylase activated by ATP
ii. Pyruvate carboxylase acitivated by Acetyl CoA
iii. Acetyl CoA carboxylase activated by Citrate
iv. Phosphofructokinase activated by AMP.
41
Clinical enzymology and biomarkers
Markers for cardiac diseases
Creatine kinase
Normal Value of CK
O
Male - 15 -100 U/l
O
Female - 10 -80 U/l
Reaction catalysed by Creatine kinase

O
CK Level starts to rise within 3 - 6 hrs of infarction.
O
Useful to detect early cases where ECG changes may be ambiguous.
O
Level not increased in hemolysis or CHF.
O
Estimation of CK(MB) isoenzyme best diagnostic marker in Myocardial infarction.
Cardiac troponins
Troponin I - released into blood within 4hrs onset of symptoms,peaks 14 - 24hrs,
o
remains elevated for 3-5 days post MI.

Troponin T - increases within 6hrs of MI,peaks 72hrs , remains elevated upto 10 - 14days.
o
Lactate dehydrogenase
Normal value - 100 - 200 U/L
:
5 isoenzymes :
i. LDH 1 - H4
ii. LDH 2 - H3 M1
iii. LDH 3- H2 M2
iv. LDH 4- H1 M3
v. LDH 5 - M4 NEET ‘ 18
Flipped LDH pattern - LDH 1 > LDH 2
seen in M.I
Time course of elevation of CK-MB and Cardiac

Troponins in Blood of M.I patients.
Markers of Myocardial infarction

42
Carbohydrates
Chemistry of Carbohydrates <
Structure of carbohydrates
Glucose
Haworth projection of
Alpha D Glucopyranose.
Ring structure formed by cyclisation is a 6 membered ring - Pyranose Ring
: Glucose exists predominantly as Pyranose ring structure - Glucopyranose
Fructose
Furan ring Haworth projection of Alpha D Fructofuranose
Ring structure formed by cyclization is a 5 membered ring - Furanose ring.

Fructose exists predominantly as Furanose ring structure- Fructofuranose.
Enediol formation
In mild alkaline solutions
0
carbohydrates containing free

sugar group ( aldehyde or
keto ) will tautomerize to form
enediols.
The interconversion of sugars through a common enediol

form is called Lowry de Bruin- Van Ekenstein transformation.
43
Monosaccharides
EPIMERS
Sugars are different from one another, only in
configuration with regard to a single carbon
atom, other than the reference carbon atom,
they are called epimers.
Example :
i. Glucose and Mannose are an epimeric
pair which differ only with respect to
C2.
Epimers of D Glucose ii. Glucose and Galactose are an epimeric
pair which differ only with respect to C4
Reactions of monosaccharides
Benedicts test
•
Detect the presence of glucose in urine.
•
Benedict's reagent contains sodium
carbonate, copper sulfate and sodium
citrate.
Any sugar with free aldehyde/ keto group
: will reduce the Benedict's reagent.

Not specific for glucose
Principle : Cu 2+ ions are reduced to Cu+ by
enediols ( Cupric to cuprous)
Reducing substances in Urine
Sugars Non-carbohydrates
i.Glucose i. Homogentisic acid
ii. Fructose ii. Salicylates
iii. Lactose iii. Vit. C ( Ascorbic acid )
iv. Galactose pentoses iv. Glucuronides
Reference : Textbook of Biochemistry for Medical Students by DM Vasudevan 7th Ed. Pg.73,316
44
Osazone Formation
@
All reducing sugars will form osazones with excess
of phenylhydrazine when kept at boiling
temperature.
@
Osazones are insoluble.
@
Each sugar will have characteristic crystal form
of osazones.
@
Osazones may be used to differentiate sugars in
Needle shaped crystals ( Glucose ) biological fluids like urine.
Needle shaped crystals ( Fructose )

Cotton ball shaped crystals ( Lactose )
Needle shaped crystals ( Mannose )

Fine long needle shaped crystals
( Xylose )
Balls with thorny edge shaped crystals

(Galactose)
Sunflower shaped crystals ( Maltose )
http://jmscr.igmpublication.org/v4-i12/14%20jmscr.pdf
45
Disaccharides
Sucrose @
a.k.a Cane sugar
@
Contains Glucose and Fructose
@
Not a reducing sugar.
@
Specific Sucrose test Positive
@
Invert sugar - hydrolysed by sucrase or
invertase products will change the
dextrorotation to levorotation.
@
Alpha 1 - Beta 2 Linkage
Lactose
Reducing disaccharide.
: Present in milk.
Beta 1 - 4 glycosidic linkage.
Maltose
:
Contains 2 glucose residues.

Alpha 1.4 linkage
: Reducing disaccharide.
Isomaltose
Reducing sugar.
: 2 glucose units combined in Alpha 1,6 linkage.
Partial hydrolysis of Starch and Glycogen produces
: isomaltose.
Oligo-1,6-glucosidase present in intestine juice can
hydrolyse isomaltose into glucose units.
46
Trehalose
Non reducing
: Major constituent of circulating fluid
(hemolymph) in insects serving as
energy storage compound.
Reference : Lehninger Principles of Biochemistry 4th edition Pg 246
Polysaccharides
STarch
Amylose
Composed of Amylose and Amylopectin.

Amylose
Soluble part ( 10-20%)
: unbranched long chain of glucose units with
o
Alpha-1,4 glycosidic linkage.
Amylopectin
/
Insoluble part.
: highly branched,branching points are made by
Alpha-1,6 linkage.
Amylopectin absorbs water forms paste like gel
Cellulose :
Most abundant organic material in nature.

Linear structure without branching.
: Beta 1,4 linkage is present.
This linkage is hydrolyzed by cellobiase.
: Herbivores and termites can digest cellulose.
47
Chitin
•
Present in exoskeletons of crustacea and insects.
•
It is composed of units of N-acetyl-glucosamine
with beta-1,4 glycosidic linkages.
Inulin
•
Long chain homoglycan composed of D-fructose units with
repeating beta-1,2 linkages.
•
It is the reserve carbohydrate present in various bulbs and
tubers, such as chicory, dahlia, dandelion, onion, garlic.
•
It is clinically used to find renal clearance value and glomerular
filtration rate.
Dextrans : highly branched homopolymers of glucose units with 1-6, 1-4 and 1-3 linkages.
Mucopolysaccharides
•
a.k.a Glycosaminoglycans (GAG).
•
are Heteropolysaccharides containing uronic acid and amino sugars (Acetylayed amino groups ,sulfate
and carboxyl groups are also present.) AIIMS ‘19
Presence of charged groups attract water molecules and produce viscous solution.
•
Excreted in abnormal amounts in Lysosomal Storage disorders.

•
Detected by 2D Gel electrophoresis techniques, CPC test,Cetavlon test and Alicante Blue staining.
•
Hyaluronic acid
•
Present in connective tissues,tendons,
synovial fluid,vitreous humor.
•
Lubricant in joint cavities.

48
Heparin
•
Repeated units of Sulfated glucosamine-
alpha-1,4-Iduronic acid.
•
Anticoagulant - activates Anti-Thrombin III
•
Present in Liver,Lungs,Spleen and Monocytes.
Chondroitin sulfate
AIIMS ‘20
•
Repeated units of D-Glucuronic acid-beta-1,3
N - Acetyl galactosamine-4 sulfate.
•
Present in cartilage,bone,tendons,cornea and
skin.
keratin sulfate Dermatan sulfate

Only GAG which doesnot contain any uronic acid.
•
Contains L- Iduronic acid and N-Acetyl
•
Repeating units - Galctose and N-Acetyl glucosamine

•
Galactosamine in Beta-1,3 linkages.
in Beta linkage. Present in Skin,Blood vessels and Heart valves
•
Present in Cornea and Tendons (Maintains corneal
:
transparency)
Reference :
1. Corneal transparency is decided by Textbook of Biochemistry for Medical Students by DM Vasudevan 7th Ed. Pg.80
A.CHONDROITIN SULPHATE C.KERATIN SULPHATE Harper’s Illustrated Biochemistry 31st Ed. Pg147
B.HYALURONIC ACID D.HEPARIN SULPHATE
Proteoglycan
Glycosaminoglycans are covalently attached to a core protein to form hybrid molecules called
Proteoglycan.
Darkfield electron micrograph
of a proteoglycan aggregate.
The proteoglycan subunits
and filamentous back- bone
are particularly well extended
in this image.
Schematic representation of a
Harper’s Illustrated Biochemistry 31st Ed. Pg599
proteoglycan complex
49
Tests for carbohydrates
Molisch test
Reagents used
1. Molisch’s reagent (alpha-naphthol in 95% ethanol)
2. Concentrated sulfuric acid.
0
General test for all carbohydrates
Carbohydrates when treated with concentrated H2SO4 undergo
: dehydration to give furfural derivatives.
These compounds condense with a-naphthol to form reddish
violet colored products (Ring)
Test is positive for formic, lactic, oxalic, citric acids and certain
:
other organic acids.
Violet coloured ring
Fehling’s Test
Fehling’s Solution Consists of:
i.Fehling’s solution A - Contains copper sulfate solution.
ii.Fehling’s solution B - Contains potassium hydroxide and
sodium potassium tartrate.
Reduction test to detect the presence of
: reducing sugars.
Carbohydrates with free aldehyde or
ketone groups reduce copper sulfate to
cuprous oxide forming yellow or brownish
red colored precipitate.
Reference : Practical Clinical Biochemistry by Dr. Shruti Mohanty

and Dr.Aparna Verma
Foulger’s Test
Foulger’s Reagent: Urea, Stannous Chloride, 40% H2SO4
: Principle: This is a specific test for ketohexoses.
A colorless solution is seen in case of glucose. Fructose gives a deep blue color solution.
: Note: Prolonged heating converts aldoses to ketoses which yield deep blue color (false
positive).
50
Seliwanoff’s test
@
Specific test for ketohexoses.
@
Concentrated HCl dehydrates ketohexoses to
form furfural derivatives which condense with
resorcinol to give cherry red color solution.
@
Prolonged heating converts aldoses to ketoses
which yield cherry red color (false positive).
Control Positive test Reference : Practical Clinical Biochemistry by Dr. Shruti Mohanty
and Dr.Aparna Verma
Bial’s Test (Tollen’s Orcinol Test)

@
Specific test for pentoses such a Ribose, Xylose
@
Bial’s reagent: Dilute solution of orcinol in 30% HCl
@
Principle: Conc. HCl acts on pentoses to form
furfurals. This condenses with orcinol to give green
color compounds.

and Dr.Aparna Verma
Rapid Furfural Test

•
Principle: Ketohexoses are rapidly converted to hydroxymethyl-furfural by HCl and form
purple colored complex with 1% alcoholic alpha naphthol.
This is a test for ketohexoses.
: There is no color in case of glucose solution, however prolonged boiling will reduce the sugar.
In case of fructose a deep purple color is produced from the moment the solution starts boiling.
51
Absorption of Carbohydrates
Glucose absorption
sglut
@
a.k.a Sodium dependent Glucose Transporter
@
2 types :
i. SGluT 1 - present in intestine
ii. SGluT 2 - present in kidney
@
Absorption from intestinal lumen into intestinal cell by
co-transport mechanism (Secondary active transport)
GLUT
@
Glucose transporter,not dependent on Sodium
@
Uniport,facilitated diffusion.
-
Glucose transporters
52
Metabolism of Carbohydrates
GLYCOLYSIS
•
a.k.a Embden Meyerhof Parnas Pathway
(EMP Pathway)
1,3- Bisphophoglycerate
Phosphoglycerate kinase
3-Phosphoglycerate
•
The enzyme common for both AIIMS ‘20
Glycolysis and Gluconeogenesis is
Phosphoglycerate kinase.
Energy yield per molecule of glucose
during Aerobic glycolysis
Steps 1,3 and 9 are key enzymes - Irreversible

Energy yield per molecule of glucose Steps 6 and 9 produce energy
during Anaerobic glycolysis Steps 5 and 10 are coupled for regeneration of NAD+
Regulatory enzymes of Glycolysis
Harper’s Illustrated Biochemistry 31st Ed. Pg159
53
Gluconeogenesis
Glucose-6-phosphate
H2O
Glucose-6-phosphatase
Pi
Glucose
NEET ‘20
•
Glucose-6-phosphatase is inhibited by Insulin.
AIIMS ‘20
Regulatory enzymes of Gluconeogenesis
PC - Pyruvate carboxylase
PEPCK - Phosphoenol pyruvate carboxykinase
Significance of Gluconeogenesis
i. Only liver can replenish sugar through
gluconeogenesis
ii. During Starvation Gluconeogenesis maintains
Blood glucose level.
iii. 6 ATP’s are required to produce 1 molecule of
Glucose
54
Malate aspartate shuttle
Oxaloacetate is generated inside the
:
mitochondria.
This oxaloacetate has to be transported
from mitochondria to cytosol, because
further reactions of gluconeogenesis are
taking place in cytosol.
This is achieved by the malate aspartate
shuttle.
Malate shuttle operates when Alanine is the
&zmsg substrate for gluconeogenesis.
8
Aspartate shuttle operates when Lactate is
the substrate for gluconeogenesis.
HMP shunt pathway
1 = Glucose 6 Phosphate Dehydrogenase

2 = Gluconolactone hydrolase
3 = 6 Phospho-gluconeogenesis dehydrogenase
4A = Isomerase
4B = Epimerase
5 = Transketolase
6 = Transaldolase
7 = Second transketolase
8 = Reversal of GLycolysis
Importance of HMP PAthway

i. To produce NADPH and pentose phosphates - ii.Ribose-5-phosphate is required for nucleic acid synthesis
•
Reductive biosynthesis so
Glucose-6-phosphate dehydrogenase deficiency
•
Free radical scavenging so
Drug induced hemolytic anemia
•
Maintain RBC membrane integrity •
Met-hemoglobinemiaThiamine deficiency leads to reduced
8
Prevention of met-hemoglobin formation transketolase activity
8
Detoxification by hydroxylation
8
Maintains transparency of lens
55
Glucose 6 phosphate deficiency
Most common enzyme deficiency seen in clinical practice.
o
X-linked recessive trait.

o
Will lead to drug-induced hemolytic anemia.

o
Drugs like
o
i.antimalarial drugs like primaquine

ii.ingestion of toxic glycosides present in fava
beans(Favism)
iii.Sulpha drugs
Enzyme deficiency offers resistance to Plasmodium infection
o
Udp glucuronic acid pathway

Importance of the Pathway
UDP Glucuronic acid (active form of
o
Glucuronic acid ) used for following

purpose :
i. Conjugation of bilirubin
ii. Conjugation of steroids
iii. Conjugation of various drugs
( more water and easily excreted )
iv. Synthesis of GAG
56
POLYOL PATHWAY OF GLUCOSE
@
Formed by reduction of glucose by
aldose reductase to sorbitol
which can then be oxidized to
fructose.
@
Sorbitol is normally present in lens
of eyes
@
In Diabetes mellitus causes
cataract.
@
Fructose in semen and CSF are all
produced by this pathway.
METABOLISM OF ALCOHOL
Alcohol Dehydrogenase (ADH) Aldehyde Dehydrogenase

NAD+ dependent cytoplasmic enzyme
@
Mitochondrial NAD+ dependent enzyme
@
Dimer and has 6 iso-enzymes.

@
Activity of alcohol dehydrogenase is more than aldehyde dehydrogenase.
Chronic Alcoholism
Manifestations
i. High NADH/NAD+ ratio v. Fatty liver and steatosis.
ii. Lactic acidosis vi. Acute attack of gout
iii. Hypoglycemia vii. CNS depression
iv. Ketogenesis viii. In pregnancy, alcohol consumption leads
to Fetal alcohol syndrome (FAS).
Laboratory Findings in Chronic Alcoholism

i. Increase in serum levels of GGT and ALT.
ii. Decrease in ADH activity is the best marker for alcohol abuse.
iii. Disialyotransferrin level in blood is a highly sensitive marker for
chronic alcohol abuse.
Liver cirrhosis
57
CORI’s cycle / lactic acid cycle
O
Lactic acid from muscle diffuses
into blood.
O
Lactate reaches liver where it is
oxidised to pyruvate.
O
Glucose regenerated can enter
blood then to muscle.
O
Process needs ATP
O
Uses :
i. Prevents Lactate accumulation
in the muscle.
ii. Reutilise lactate from muscle
and RBC for gluconeogenesis.
Pasteur effect : under anaerobic condition glycolysis is inhibited.
Warburg hypothesis :
Cancer cells utilise energy from glycolysis and they require less oxygen than their normal counterparts.
cahill cycle / Glucose alanine cycle

Alanine is transported to liver,transaminated
o
to pyruvate and converted to glucose.

Important during conditions of starvation.
o
Uses : o
i. Carries amino group to the liver

ii. Alanine as a substrate for
gluconeogenesis during starvation.
58
Fructose metabolism
Entry of Fructose into Glycolysis
Hereditary fructose intolerance

om
Autosomal Recessive
om
Biochemical defect : Deficiency of Aldolase B
or
Asymptomatic until Fructose or sucrose is ingested.
or
Jaundice, Hepatomegaly
or
Vomiting, Lethargy, Irritability ,Convulsions and Hypoglycemia
Lab Diagnosis
Rapid Furfural test and Seliwanoff’s test Positive
or
Treatment
or
Lactose free Diet.
glycogen metabolism
or
Glycogen phosphorylase removes glucose
as glucose- 1-phosphate from glycogen
(phosphorolysis).
or
It contains pyridoxal phosphate (PLP) as
a prosthetic group.
or
If glycogen phosphorylase alone acts on a
glycogen molecule, the final product is a
highly branched molecule; it is called limit
dextrin.
59
Relative changes in plasma hormones and metabolic
fuels during the onset of starvation.
Glycogen storage disorders

60
Mc Ardle’s Disease
AIIMS ‘18
O
a.k.a Type V Glycogen Storage Disorder.
O
Caused by deficiency of Muscle Phosphorylase activity.
Symptoms
Generalised Muscle weakness.
O
Hypotonia.
O
Excercise intolerance with muscle cramps and pain.

O
“ Second Wind “ phenomenon is seen.

O
Muscle biopsy showing vacuoles myopathy.

Investigations
i. Serum Creatine kinase level elevated after excercise.
ii. Increased level of blood ammonia,ionosine,hypoxanthine and
uric acid.
iii. Muscle biopsy
iv. Phosphorus MRI
Treatment
Avoid strenuous excercise
O
Glucose or Sucrose given before excercise.

O
Burgundy coloured urine after excercise -

High Protein diet
O
Excercise induced Myoglobinuria 2 to Vit B6 supplementation

O
Rhabdomyolysis
61
Von Gierke Disease
AIIMS ‘17
•
a.k.a Type I Glycogen Storage Disorder
Glucose-6-Phosphatase deficiency ( Type Ia)
Glucose-6-Phosphate translocase deficiency ( Type Ib)
Clinical manifestations
Seizures ( Fasting Hypoglycemia )
•
Doll Like facies ( fat cheeks relatively thin extremity ,short stature)
•
Protruberant abdomen - Hepatomegaly, Enlarged kidney.

•
Delayed puberty - Fertility normal

•
Biochemical hallmarks
•
Hypoglycemia
•
Lactic acidosis
•
Hyperuricemia
Protuberant abdomen
•
Hyperlipidemia - Plasma appears “ Milky”
Treatment
•
Continuous nasogastric infusion of glucose or oral

administration of uncooked cornstarch.
•
Sucrose,Fructose and Sorbitol limited or restricted diet.
•
Calcium and Vit D Supplementation
•
Orthotopic liver transplantation is a potential cure.
Milky Plasma Normal Plasma Reference : Textbook of Biochemistry for Medical Students by DM Vasudevan 7th Ed. Pg 129
62
Pompe disease
@
a.k.a Type II Glycogen Storage Disorder
@
Deficiency of Acid Alpha -1,4 glucosidase responsible
for degradation of glycogen in lysosomes.
@
Autosomal Recessive
@
Gene for for the enzyme is on Chromosome 17.
Infantile Pompe disease
i. Floppy infant - hypotonia
ii. Neuropathic bulbar weakness
iii. Feeding difficulties
iv. Macroglossia
Late Onset Pompe Disease v. Hepatomegaly
Diagnosis vi.Hypertrophic cardiomyopathy f/b death
i. Elevated Serum Creatine kinase Late onset pompe disease
ii. Elevated Aspartate amino transferase i. Slowly progressive proximal muscle weakness
iii. Elevated levels of Lactate dehydrogenase with truncal involvement.
iv. Enzyme Assay - deficient Alpha Glucosidase ii. Pelvic girdle,paraspinal muscles and diaphragm
v. ECG - High voltage QRS complex and are muscle groups seriously affected.
shortened PR interval. iii. Ptosis
vi.Elevated urinary glucose tetrasaccharides iv. Lingual weakness.
Chest X ray and Muscle histology findings

Before Treatment AfterTreatment
Treatment
Supportive or Palliative
@
Specific enzyme replacement therapy

@
with recombinant human acid alpha

glucosidase.
High protein diet and excercise.
@
Nocturnal Ventilatory Support as when

@
needed.
63
Mucopolysaccharidosis
Hurler syndrome
Mucopolysaccharidoses I
i Mutation of IDUA gene on Chr. 4
Deficient Alpha-L-Iduronidase
: Normal at birth
Radiographic findings
Corneal clouding, Large tongue.
Inferior portions of ilia are •

Metacarpals and phalanges are
hypoplastic and shallow abnormal short.
acetabular fossa. •
Bullet shaped phalanges
Clinical manifestations Treatment

Hepatomegaly Prominent forehead Enzyme replacement using
•
: Coarse facial features

•
Joint stiffness
• Recombinant Alpha-L-
•
Corneal clouding Short stature
• Iduronidase(Aldurazyme)
Large tongue Skeletal dysplasia Bone Marrow Transplantation
@
:
•
Abnormal spacing of teeth with dentigerous cyst.

Progressive ventricular enlargement - communicating
hydrocephalus.
Recurrent URTI and ear infections Inguinal hernia
0
: Persistent noisy breathing
# Earliest radio graphic signs - Thick ribs and ovoid vertebrae.

Reference : Nelson Textbook of Pediatrics 20th Ed. Pg 737
64
Hunter disease
Mucopolysaccharidosis II
@
X Linked Recessive disorder

O
Deficient Iduronate 2 sulfatase

O
Coarse facial features
O
Short stature
O
Dysostosis complex
O
Joint stiffness
O
Coarse facial features,Joint stiffness Intellectual disability

O
Chronic diarrhoea
O
Treatment
Enzymes replacement - Elaprase
O
Communicating hydrocephalus
O
Spastic paraplegia
O
# Lack corneal clouding

https://www.semanticscholar.org/paper/Severe-Hunter-Syndrome-(Mucopolysaccharidosis-II)-X-Burruss-Wood/f0bae57c38e5e6b2ad1b6ce2b1cf3aa8076e259b
Sanfilippo disease
Autosomal Recessive
O
4 types : O
i. Sanfilippo Disease A - deficient Heparan sulfamidase

ii. Sanfilippo Disease B - deficient N-Acetyl alpha D glucosaminidase
iii. Sanfilippo Disease C - deficient Acetyl CoA Alpha glucosaminide N-
Acetyl transferase
iv. Sanfilippo Disease D - deficient N- Acetylglucosamine-6-sulfatase.
Delayed development
O
Hyperactivity with aggressive behaviour

O
Coarse hair and hirsutism

O
Sleep disorders
O
Hepatosplenomegaly # Clear corneas

O

65
Morquio Disease
@
Autosomal Recessive
@
2 types :
i. Morquio A - deficient N-Acetyl galactosamine-6-sulfatase
ii. Morquio B - deficient Beta Galactosidase
@
Defective degradation of Keratan sulfate
CLinical Manifestations
Short trunk,Dwarfisim
@
Small teeth with
0
Fine corneal deposits

@
abnormally thin enamel.
Skeletal dysplasia
@
Frequent caries formation.
@
Preservation of intelligence
0
Hepatomegaly @
Waddling gait
@
Life threatening@
Kyphosis
@
Atlantoaxial instability.
Radiographic findings
Platyspondyly with Proximal pointing Delayed ossification of

maintained intervertebral of metacarpals femoral heads.
disc and central breaking Metaphysical widening
in Proximal femur
https://www.researchgate.net/figure/Morquios-syndrome-Radiographs-of-spine-A-B-show-platyspondyly-with-maintained_fig6_267742738
66
Maroteaux-Lamy disease
0
Mucoploysaccharidosis VI
Mutation in ARSB gene on Chr. 5 codes
: Deficient N-Acetylgalactosamine-4-sulfatase(Aryl sulfatase B)
clinical manifestations
Corneal clouding
0
Communicating hydrocephalus
0
Coarse facial features Dysostosis multiplex
:
0
Joint stiffness
Treatment
Enzyme replacement therapy - Naglazyme
Corneal clouding
Bullet-shaped metacarpal bones and phalanges. Reference : Nelson Textbook of Pediatrics 20th Ed. Pg 742
Slanting/ convergent distal radius and ulna. https://radiopaedia.org/cases/mucopolysaccharidosis-type-vi-maroteaux-lamy-syndrome-1?lang=gb
Sly syndrome
Mucopolysaccharidosis VII
:Mutation in gene GUSB gene on Chr.7

Deficiency of Beta-Glucuronidase
Most severe form presents as Non Immune Fetal Hydrops Fetalis
•
Thick skin Dysostosis complex
0
•
Normal intelligence Corneal clouding varies
0
•
Visceromegaly
67
Metabolism of galactose
Galactosemia
NEET ‘ 18
•
Autosomal Recessive
•
Deficiency of Galactose-1-phosphate uridyl transferase
•
Jaundice Poor weight gain
•
•
Hepatomegaly Oil drop Cataract
•
•
Vomiting Intellectual disability
•
•
Hypoglycemia Hypergonadotrophic hypogonadism s/i 80-90%
•
Oil Drop Cataract due to •

Irritability Female patients.
accumulation of Dulcitol
Investigations
Hyperbilirubinemia - unconjugated
•
Blood - Galactosemia
•
Urine - Galactosuria
•
HPLC - detect levels of Galactose-1-

•
phosphate transferase in RBC.

Treatment
Lactose free diet
•
68
Lipids
Chemistry of Lipids
Polyunsaturated fatty acids
so
PUFA exist in cis configuration in naturally occuring lipids.
Clinical significance
Linoleic and Linolenic acids are Essential Fatty Acids
so
Arachidonic acid is precursor of Prostaglandins

so
Pentatonic acid ( Omega 3 unsaturated fatty acid )present in fish oils is of nutritional importance.
so
Eicosanoids are derived from 20 carbon Arachidonic acids.

so
Phosphatidylcholine or lecithin
Structure of Lecithin
Action of Phospholipases
1. Phospholipase A 1 - products formed are Lysolecithin and fatty acid
2. Phospholipase A 2 - products formed are Acylglycerophosphorylcholine and Fatty acid
3. Phospholipase C - products formed are 1,2 Diacylglycerol and Phosphoryl choline
4. Phospholipase D - products formed are Phosphatidic acid and cholime
69
Respiratory distress syndrome
It is due to defect in the biosynthesis of Dipalmitoyl lecithin.
so
It is the main surfactant.

so
Premature infants don’t synthesize enough DPL

so
Cephalin
so
a.k.a Phosphatidylethanolamine
so
Ethanolamine is present instead
of choline.
so
Found in biomembranes.
Phosphatidylinositol
so
Phosphatidyl inositol bisphosphate or PIP2
is present biomembranes.
so
Act as second messenger.
Sphingomyelin
Are the only sphingolipid that contain phosphate and have no sugar moiety.
: Found in large quantities in nervous system
Can act as emulsifying agent and detergent.
so
Combine with fatty acid forms Ceramide ( component of Glycophospholipids )
70
Plasmalogen
@
Phospholipids which have an aliphatic
long chain alpha and beta unsaturated
alcohol.
so
Found in Biomembranes in brain and
muscle.
Fatty acid metabolism

Structure of Micelle
so
Formed by 2 monoacyl- glycerols, long chain
fatty acids, cholesterol, phospholipids and
lysophospholipids are incorporated into
molecular aggregates to form mixed micelle.
so
Spherical particles with a hydrophilic exterior
and hydrophobic interior core.
so
Essential for the absorption of fat-soluble
vitamins such as vitamin A, D and K.
Absorption of fatty acids

so
Long chain fatty acids are absorbed into
the intestinal cell wall, where they are
re-esterified, made into chylomicrons
and enter into lymphatics.
so
Short chain fatty acids are directly
absorbed into blood capillaries. Bile
acids are reabsorbed into portal vein.
71
Abnormalities in absorption of lipids
i. Defective absorption - Steatorrhea daily excretion more than 6g per day.
ii.Defective absorption may be due to :
a.Celiac disease
b.Surgical removal of intestine
c.Obstruction of bile duct
iii.Chyluria - urine appears milky due to lipids
iv.Chylothorax
Beta oxidation of fatty acids
so
Palmitic acid (16 C) needs 7 cycles of
Beta oxidation.
so
8 molecules of Acetyl Coa is released.
so
Energetics of Beta Oxidation :
8 Acetyl CoA × 10 = 80 ATP
7 FADH2 × 1.5 = 10.5 ATP
7 NADH × 2.5 = 17.5 ATP
Gross Total = 108 ATP
Net yield = 108 - 2 = 106 ATP
Oxidation of Odd chain Fatty acids produce Propionyl CoA (3C)

72
so
As a general rule, fatty acids cannot be used for gluconeogenesis.
so
But 3 carbon units from Odd chain fatty acids are Gluconeogenic.
so
Propionate is Gluconeogenic.
Role of carnitine in transport of acyl groups

Carnitine is Beta Hydroxy Gamma 0
Trimethyl Ammonium Butyrate.

Synthesised from Lysine and Methionine
O
in Liver and Kidney.

Vit. C is essential for the synthesis of
O
Carnitine.
It is involved in transfer of activated
O
fatty acids from cytoplasm to

mitochondria.
Structure of co-enzyme A
Organic acidurias
O
Disorders of metabolism of Fatty acids,Branched chain and Aromatic Amino Acids and Citric acid cycle.
O
Characterized by accumulation of organic acids in the body tissues and their excretion in urine.
O
Some important ones are :
73
Fate of Propionyl CoA
0
Propionyl CoA is derived from
i. Oxidation of Fatty acids
ii. Metabolism of Valine
iii. Metabolism of Isoleucine.
ALPHA OXIDATION of fatty acids

Fatty acids are oxidized by removing carbon atoms, one at a time,from the carboxyl end.
•
Important in Brain occurs in Endoplasmic Reticulum.

•
No need for FA activation.

•
Does not generate Energy.

•
Refsum's Disease
Metabolic error due to lack of alpha-hydroxylase (phytanic acid oxidase).
:
caused by defects in PEX7.
Alpha oxidation doesnot occur.
clinical Features
Neurological symptoms
: Polyneuropathy
Retinitis pigmentosa
: Nerve deafness
Cerebellar ataxia Retinitis pigmentosa
:
Infantile Refsum's Disease
It is a peroxisomal disorder.
Phytanic acid accumulates along with VLCFA
74
DE NOVO SYNTHESIS OF FATTY ACIDS
a.k.a Lynen's spiral.
: Extramitochondrial or cytoplasmic
fatty acid synthase system.
Major fatty acid synthesized de novo is
: palmitic acid, the 16C saturated fatty

acid.
Occurs in liver, adipose tissue, kidney,
brain, and mammary glands.
75
Ketogenesis
Acetoacetate is the primary ketone body.
: Betahydroxy butyrate and acetone are
secondary ketone bodies.
Synthesized in liver mitochondria.
: Beta hydroxyl butyrate does not contain a

keto group,it contains alcohol group.
Almost all tissues and cell types can use
ketone bodies as fuel, with the exception of
liver and RBC.
I
Acetoacetate is activated to Acetoacetyl CoA
by thiophorase enzyme.
Ketosis
Causes :
i. Diabetes mellitus
ii. Starvation
iii. Hyperemesis in Early pregnancy
Features of Ketosis
i. Metabolic acidosis v. Osmotic diuresis
ii. Reduced buffers vi.Sodium loss
iii. Kussmaul’s respiration vii.Dehydration
iv. Smell of Acetone in breath viii.Coma
Diagnosis
i. Rothera’s test.
76
Rothera’s test
•
Picture shown is that of Positive Rothera’s test.
•
There will be formation of violet ring at the junction of 2 liquids.
•
Positive test indicates presence of Acetone,Acetoacetic acid.
•
Not answered by Beta hydroxy butyrate. AIIMS ‘17
•
Ketone bodies are present in urine during starvation state.
Rothera’s test Reference : Textbook of Biochemistry for Medical Students by DM Vasudevan 7th Ed. Pg 429
ZELLWEGER SYNDROME
O
a.k.a Cerebrohepatorenal syndrome NEET ‘19 AIIMS ‘19 AIIMS ‘18
O
Autosomal Recessive
O
Its a peroxisomal biogenesis disorder ( Failure to import 1 or more
proteins into organelle).
O
There is defective oxidation of VLCFA.
O
There is accumulation of C 26 - C 38 polygenic acid in brain.
Clinical Features
Typical facial appearance ( high forehead,hypo plastic
O
supraorbital ridges and epicanthic folds)

Severe weakness
O
Hypotonia
O
Neonatal seizures
O
Eye abnormalities
O
Diagnosis
Demonstration of abnormally high levels of VLCFA’s in Plasma
O
# X - linked Adrenoleukodystrophy (ALD) is the most common peroxisomal disorder,rest all

peroxisomal disorders are AR.
77
Cholesterol and Lipoproteins

Structure of Lanosterol and cholesterol
electrophoretic and ultra-centrifuge patterns of lipoproteins

@
Oil Red O stain is used in
Electrophoresis.
@
Lipoproteins with higher protein
content will move faster towards
the anode and those with
proteins have minimum mobility.
78
Comparison of sizes of lipoproteins
Structure of chylomicrons
•
Formed in the intestinal mucosal
cells, and secreted into the
lacteals of lymphatic system.
•
Rich in Triglyceride.
•
Apo-C and Apo-E are added from
HDL in blood during transport.
Largest of Plasma Lipoproteins.
:
Metabolism of chylomicrons
Chylo - Chylomicrons
Ch - Cholesterol
TG - Triglyceride
: A,B48,C-II and E are Apoproteins
79
Lipoprotein
Are compound lipids formed as a combination of
: proteins and lipids.
Consist of non polar core and a single surface
layer of amphipathic lipids.
Non polar core consists of TAG and Cholesteryl
:
ester.
Low density lipoprotein

Transports cholesterol from liver to
peripheral tissues.
only apoprotein present in LDL is Apo B100.
: Mostly derived from VLDL.
Half life of LDL in blood is about 2 days.
:
Uptake and fate of ldl

Function of LDL
About 75% of the plasma cholesterol is
incorporated into the LDL particles.
Transports cholesterol from liver to the
peripheral tissues.
Reference : Textbook of Biochemistry for Medical Students by DM Vasudevan 7th Ed.

80
Lipoprotein (A)
☐
Is strongly associated with
Myocardial Infarction.
a.k.a Little Rascal
☐
Attached to Apo B-100 by
disulphides bond.
☐
It interferes with plasminogen
activation and impairs
fibrinolysis,leading to unopposed
intravascular thrombosis.
HIGH DENSITY LIPOPROTEIN

☐
Main transport form of cholesterol from peripheral tissue to liver
This is called reverse cholesterol transport by HDL.
:Only excretory route of cholesterol from the body is the bile.
PUFA is anti-atherogenic.
Bile acids
Structure of Bile acids
•
Synthesized in the liver from
cholesterol.
They contain 24 carbon atoms.
: All of them have an alpha-
oriented (projecting below the
plane of ring) hydroxyl group at
position 7.
81
Formation of bile salts
Rate limiting step is
: introduction of hydroxyl group

by enzyme 7 Alpha hydroxylase.
This is the first step in
formation of bile salts.
Primary Bile Acids

i. Cholic acid
ii. Chenodeoxycholic acid
Secondary Cholic Acids

i. Deoxycholic acid
ii. Lithocholic acid
82
Lipid Storage Disorders

Gauchers disease
Enzyme defect : Beta - glucosidase
: Lipid accumulating - Glucocerebroside
Clinical features
Bruising from thrombocytopenia
: Chronic fatigue secondary to anemia
hepatomegaly with or without elevated
liver function test results
Splenomegaly
: Bone pain
Juvenile hepatosplenomegaly
Gaucher cell in the

reticuloendothelial system,
particularly in the bone marrow.
“Wrinkled paper appearance“
Nelson Textbook of Pediatrics 30th Ed Pg709
Niemann pick disease

Enzyme defect : Sphingomyelinase
Lipid Accumulating - Sphingomyelin
Clinical features
Severe CNS damage
: Mental retardation,
Hepatosplenomegaly
: Cherry red spot in macula.
Death occurs by 2 years of age Cherry red spot in macula.
83
Krabbe’s leukodystrophy
a.k.a Globoid cell leukodystrophy
0
a- ' ,
y
l
Autosomal Recessive fatal disorder
o
I
t
'
,
l -
l
Enzyme defect :Beta - galactosidase
•
/
1- Lipid Accumulating : Galactocerebroside
•
-
l
/ -
l - '
y l
l
"
1
'
I
,
I
,
,
"
l
"
Clinical features
Severe mental retardation.
/ -
,
y - / L -
1-
.
I ,
' '
n .
Total absence of myelin in CNS

: Globoid bodies in white matter
-
s
l
l
l
l l
Histology of brain tissue shows many large multinucleated

(“globoid”) cells of 20–50 μm in diameter in the
neighborhood of a blood vessel.
Fabry’s disease
•
X-linked inborn error of glycosphingolipid metabolism
caused by the absent or markedly deficient activity of
α-galactosidase A (α-gal A).
•
Lipid accumulating : Ceramide trihexoside
(Kidney is the site of accumulation)
Clinical features
Angiokeratomas
•
Corneal opacities
Angiokeratomas
•
Abdominal pain
•
Fabry crises, lasting from minutes to several days

•
Treatment
•
Phenytoin and/ or carbamazepine
Polarised light microscopy of urine showing

“Maltese Cross” pattern. Reference : Textbook of Biochemistry for Medical Students by DM Vasudevan 7th Ed. Pg 193
Nelson Textbook of Pediatrics 30th Ed Pg 712
84
Tay Sachs disease
•
Enzyme defect : Hexosaminidase A
•
Lipid accumulating : Ganglioside (GM2)
Clinical Features
Mental retardation.
•
Cherry red spot in the macula.

•
Progressive deterioration.
•
Death by 3–4 years

•
Cherry Red spot in Macula

85
Molecular Genetics
Structure of D N A
Chargaff’s Rule
NEET ‘20
•
a.k.a Base Pairing Rule
•
States that number of purine is always equal to number of

pyrimidines.
•
Adenine will always pair with Thymine and Cytosine will always
pair with Guanine in DNA.
types of bonds in DNA

Phosphodiester bond
Deoxyribionucleotides are linked
•
together by phosphodiester bonds to

form polynucleotides
AIIMS ‘17
Hydrogen bond
DNA strands are held together by
•
Hydrogen bonds.
2 hydrogen bonds between A and T
•
3 hydrogen bonds between C and G

•
86
Nucleosome
•
Nuclceosomes contains 4 major types of histones :

i.H2A form dimers
ii.H2B
I
iii.H 3 form tetramer
iv.H 4
•
These 4 core histones are subjected to at least 6 types
of post translational modifications( PTM).
•
H1 histones are the least tightly bound to chromatin.
•
Histones are proteins containing higher concentrations
Nucleosome
of Basic amino acids.
1.75 turns of superhelical DNA is wrapped around
i
histone octamer,protecting 145- 150bp of DNA and
forming nucleosome core particle.
DNA condensation
DNA Double Helix 2 nm in diameter
Nucleosomes 10 nm in diameter
Chromatin fibril 30 nm in diameter
t
Extended Chromosome 300 nm in diameter
Chromosome 1400 nm in diameter
87
DNA Replication
Semiconservative synthesis
DNA replication starts with recognition of

site of origin of replication (ORI) recognized
by origin recognition complex (ORC).
DNA helicase unwinds the DNA
DNA replicase system/ replisomes:

i. The protein A or DnaA binds at specific sites of origin, and opens the duplex.
ii. The relief of supercoil needs topoisomerases.
iii. Helicase or DnaB separates the strands, using ATP. Helicase moves in both
directions, separating the strands in advance for replication, forms a replication
bubble with two replication forks.
iv. Single stranded DNA binding proteins (SSB) stabilise the complex.
Each replication fork contains:

a. DNA helicase which unwinds a short region of DNA helix
b. Primase that initiates synthesis of a short RNA segment
c. DNA polymerase enzyme that synthesizes the new strand
of DNA
d. DNA is prevented from rewinding by SSBs.
RNA primer, about 100–200 nucleotides long

synthesized by the RNA primase.
RNA primer is removed by DNAP, using
exonuclease activity and is replaced with
deoxyribonucleotides by DNAP
Using DNA polymerase, the 3′ hydroxyl group of the
end nucleotide combines with the 5′ phosphate group
1 of the new deoxynucleotide. The pyrophosphate is

released from the deoxynucleoside triphosphate
88
DNA polymerase carries out the

sequential addition of each nucleotide
complementary to the one in the Polymerization of the new strand
template strand. of DNA is taking place from 5′
to 3′ direction, template is read
in the 3′ to 5′ direction
DNA synthesis is always in the 5′ to 3′

direction in both strands. The strand which is
discontinuously synthesized is referred to as
the “lagging strand” otherwise called
“retrograde strand” and the one continuously
polymerized as the “leading strand”
The small DNA molecules attached to its own primer RNA are called
Okazaki fragments.. The synthesis along the lagging strand is in 5′ to 3′
direction. As it moves, the primase synthesizes short RNA primer, to
which deoxyribonucleotides are added by DNA polymerase. (DNAP-alpha
synthesizes short pieces, which are extended by DNAP-delta. This delta
enzyme has a role in the synthesis of both leading and lagging strands).
DNAP-alpha has primase activity.
DNA synthesis continues until the primer and previously

added Okazaki fragment is encountered. Then the RNA
primer is removed by DNAP, using exonuclease activity
and is replaced with deoxyribonucleotides by DNAP. The
remaining nick is sealed by the DNA ligase.
Condensation into Chromatin Structure
Reference : Textbook of Biochemistry for Medical Students by DM Vasudevan 7th Ed. Pg 579 - 581
89
Major Proteins Involved in DNA Replication

DNA polymerases Add nucleotides to a strand growing 5′ to 3′, copying a DNA
template 3’ to 5’
Primase Synthesizes RNA primers
Helicases Separate parental DNA strands, i.e., unwind the double
helix
Single-strand Prevent single strands of DNA from reassociating
binding proteins
Topoisomerases Relieve torsional strain on parental duplex caused by
unwinding
Enzymes that RNase H hydrolyzes RNA of DNA–RNA hybrids
remove primers
Flap endonuclease 1 (FEN1) recognizes flap
(Unannealed portion of RNA) near 5’-end of primer
and cleaves downstream in DNA region of primer; the
flap is created by polymerase δ displacing the primer
as the Okazaki fragment is synthesized
DNA ligase Joins, by forming a phosphodiester bond, two
adjacent DNA strands that are bound to the same
template
PCNA Enhances processivity of the DNA polymerases; binds
to many proteins present at the replication fork
90
Telomere and Telomerase
INICET’2O
i. Replication takes place from 5′ to 3′
direction in the new strand.
ii. DNA polymerase enzyme is not able to
synthesize the new strand at the 5′ end
of the new strand.
iii. A small portion in the 3′ ends of the
parent strands could not be replicated
iv. This end piece of the chromosome is
called telomere.
v. Enzyme, telomere terminal transferase
or telomerase helps in replication of the
end piece of chromosomes (noncoding
repetitive sequences).
vi. The small portion left; is degraded by
exonucleases.
vii. Could cause aberrant recombinations
or end to end fusions.
viii. If no mechanism to replicate
Covered by many copies of shelterin
telomeres occurs, length of the
chromosomes keeps reducing after each
cell division, stability of the chromosome
is lost.
ix. The shortening of telomere end is
prevented by enzyme telomere terminal
transferase or telomerase, contains an
RNA component, which provides the
template for telomeric repeat synthesis.
x. Telomerase acts like a reverse transcriptase. Telomerase recognises 3′ end of telomere, and then
a small DNA strand is synthesized.
xi. Telomeric DNA is covered by shelterin protein complexes that specifically bind to telomeric
DNA repeats and protects telomeres from being recognised by DNA repair proteins, thereby
preventing fusion of chromosomes.
Netters Essential Biochemistry 1st edition, pg 26
91
Human Diseases of DNA Damage Repair
Defective Nonhomologous End-Joining Repair (NHEJ)

1. Severe combined immunodeficiency disease (SCID)
2. Radiation-sensitive severe combined immunodeficiency disease
(RS-SCID)
Defective Homologous Repair (HR)
1. AT-like disorder 4. Werner syndrome (WS)
(ATLD) 5. Rothmund-Thomson syndrome (RTS)
2. Nijmegen breakage 6. Breast cancer susceptibility 1
syndrome (NBS) and 2 (BRCA1, BRCA2)
3. Bloom syndrome (BS)
Defective DNA Nucleotide Excision Repair (NER)

1. Xeroderma pigmentosum (XP)
2. Cockayne syndrome (CS)
3. Trichothiodystrophy (TTD)
Defective DNA Base Excision Repair (BER)
1. MUTYH-associated polyposis (MAP)
Defective DNA Mismatch Repair (MMR)

1. Hereditary nonpolyposis colorectal cancer (HNPCC)
Reference : Harper’s Illustrated Biochemistry 31st Ed. Pg 370
92
Different Classes of R N A
Transfer rna
D
a.k.a t RNA AIIMS ‘18
D
They transfer amino acids from cytoplasm to
ribosomal protein synthesising machinery.
D
Easily soluble hence soluble RNA or sRNA.
so
73 - 93 nucleotides in length.
Acceptor Arm
so
Is at 3’ End
so
7 base pairs.
so
Forms an ester bond with carboxyl end of AA
DHU Arm
so
Dihydrouridine is present in that area.
so
Serves as recognition site for enzyme
which adds AA
Anticodon Arm Pseudouridine Arm
so
recognizes triplet nucleotide codon. so
Contains pseudouridine
so
Specificity of tRNA resides in anticodon site. so
involved in binding to ribosomes.
if short extra arm is present (3-5 base pairs ) Class I tRNA

# Only 75% tRNA contains extra arm
if long extra arm is present (13 - 21 base pairs) Class II tRNA
93
Epigenetics
INICET’2O AIIMS’18 NEET’18
- Types of epigenetic modifications:
:
(A) Histones can undergo phosphorylation (Ph), methylation (Me), and acetylation of specific
lysine residues (Ac), these modifications are involved in chromatin remodeling and transcriptional
regulation.
- Methylation at the CG sequence of the promoters transcriptionally inactivates chromatin.
- Methylation occurs naturally on cytosine bases at CpG sequences and is involved in controlling
the correct expression of genes.
(B) DNA molecules are methylated by the addition of a methyl group to carbon position 5 on
cytosine bases, a reaction catalyzed by DNA methyltransferase enzymes, which maintains repressed
gene activity.
- Associated with triggering histone deacetylation, chromatin condensation, and gene silencing.
(C) mRNA is translated into a protein product, but this process can be repressed by binding of
microRNAs (miRNA), a class of noncoding RNA (ncRNA).
- Epigenetic states can be modified by environmental factors which may result in the expression of
abnormal phenotypes. These epigenetic modifications control gene expression and changes are also
inherited. Reference : https://journals.plos.org/plospathogens/article/figure?id=10.1371/journal.ppat.1003007.g004
Textbook of Biochemistry for Medical Students by DM Vasudevan 7th Ed. Pg 623
94
Recombinant DNA Technology AIIMS’19
i. Restriction enzymes cleave specific
sequences of DNA
ii. Obtain DNA fragments for study/
÷
insertion into the DNA from donor.
iii. Chimeric or recombinant DNA obtained
iv. Hybridisation (DNA strands can base-
pair with complementary strands of DNA
or RNA).
v. Labeled DNA used as a probe to identify
homologous (complementary sequences)
DNA or RNA.
vi. Gel electrophoresis separates DNA
fragments by size.
vii. Nucleotide sequence of DNA
determined. used to deduce the amino acid
sequence of the protein produced from
the DNA.
viii. Large quantities of DNA produced by
the PCR
ix. Fragments of DNA obtained from genomic DNA or DNA copied from mRNA (cDNA) amplified by
PCR and cloned (inserted into another organism, foreign DNA replicated and expressed).
x. Large quantities of the protein product obtained.
Specificity of restriction enzymes (The arrows show the

site or cut by the enzyme)
95
obtaining fragments of DNA

i. Short sequences of DNA (oligonucleotides) synthesised in vitro
ii. Are used as primers for DNA synthesis or as probes for the detection of DNA or
RNA sequences.
iii. Restriction endonucleases (RE) cleave DNA into fragments. AIIMS’19 AIIMS’17
a. Restriction endonucleases recognize short sequences in DNA (palindromes) and cleave

both the strands within this region.
(1) The enzyme EcoR1 (example for RE)
cleaves a region between an A and a G on
each strand, generating two products.
(2) The single-stranded regions of the products
allow them to reanneal or to recombine
with other DNA that has been cleaved by
the same restriction endonuclease.
b. A DNA fragment containing a specific gene is isolated from the cellular
genome with restriction enzymes.
(Genes isolated from eukaryotics usually contain introns, bacteria does not)
iv. mRNA for a gene can be isolated, and a DNA copy (cDNA) can be produced by
reverse transcriptase.
v. cDNA does not contain introns, nor does it contain the promoter region of a
gene, as that region is not transcribed.
96
CRISPR-Cas 9
i. Clustered regularly interspersed short palindromic repeats -associated gene 9

ii. Found in many bacteria
iii. Form of acquired, or adaptive immunity to prevent reinfection of a bacterium by specific
bacteriophages.
iv. CRISPR uses RNA-based targeting to bring the Cas9 nuclease to foreign (or any complementary)
DNA.
v. Within bacteria this CRISPR-RNA-Cas9 complex degrades and inactivates the targeted DNA.
vi. CRISPR system adapted for use in eukaryotic cells, is shown to be an RNA-directed site-specific
nuclease
vii. Highly efficient, and very specific technology for the manipulation of DNA and genetic analysis of
mammalian cells
- CRISPR-Cas9 mediated gene repair occur via nonhomologous end joining

\ AIIMS’19
(NHEJ) mechanism.
Reference : DM Vasudevan Textbook of Biochemistry 7th edition, Pg 626

97
Blotting
i. Southern blot transfer

- used for DNA
ii. Northern blot transfer
- used for RNA
iii. Western blot
- used for proteins
iv. Southwestern blotting
- Also known as overlay blot
- Examines protein–nucleic acid
interactions or protein–protein
interactions
98
Amplification and Hybridisation Techniques

POLYMERASE CHAIN REACTION
INICET’2O
- Method of amplifying
/
target sequence of DNA.
Denaturation
- Seperation of DNA by melting
through 95 °C for 15sec to 2min
- Converts double stranded DNA to
single stranded DNA
Annealing
- Hybridization of two oligonucleotides
used as primers to the target DNA
- At 3’ end, cooling to 50 °C for 0.5
to 2 min
Extension
- Elongation/ polymerase reaction
of DNA chain by nucleotide
addition from the primers
(new DNA strand synthesised)
- Using DNA polymerase/ Taq
polymerase as catalyst in the
presence of Mg2+ ions, dNTPs.
- At 72 °C for 30 sec
AIIMS’18
Q: RT PCR is ............. based technique.
- RNA
99
RT-PCR
•
Reverse transcription polymerase chain
reaction (RT-PCR) is a laboratory
technique combining reverse transcription
of RNA into DNA (in this context called
complementary DNA or cDNA) and
amplification of specific DNA targets using
polymerase chain reaction (PCR).
•
It is primarily used to measure the amount
of a specific RNA.
•
This is achieved by monitoring the
amplification reaction using fluorescence, a
technique called real-time PCR or
quantitative PCR (qPCR).
•
Combined RT-PCR and qPCR are routinely
used for analysis of gene expression and
quantification of viral RNA in research and
clinical settings.
3.RNA is used in which of the following techniques ?

A.Sanger’s Technique
B.Western Blot
C.RT-PCR
D.Southern Blot [INICET 2021]
Southern blot technique

•
This is used to detect a specific segment of
DNA in the whole genome.
1.DNA is cut with restriction enzymes;
Electrophoresis on agar gel.
2.DNA fragments blotted on nitrocellulose
membrane
C3.DNA probe added to membrane
4.Bands visualised by autoradiography
100
Comparison of blot transfer techniques
Sanger sequencing
• Sanger sequencing is a method of DNA sequencing based on the selective incorporation of chain-
terminating dideoxynucleotides by DNA polymerase during in vitro DNA replication.
101
Patterns of Inheritance
Autosomal dominant inheritance
•
Phenotypic expression of the disease, even if
one allele is abnormal or in heterozygous state.
•
Trait may be transmitted by either parent and
can affect both male and female offsprings.
Examples :
i. Marfan’s syndrome. NEET ‘19
ii.Achondroplasia ( Dwarfisim).
iii. Hereditary Spherocytosis.
iv. Osteogensesis imperfecta
v. Neurofibromatosis 1 nad 2
vi. Hypercholesterolemia
vii. Huntington’s disease
viii. Spinocerebellar Ataxia
ix. Hutchinson-Gliford Progeria
x. Noonan syndrome
Normal Affected
Marfans’s Syndrome
Achondroplasia
Reference : Harrison’s Internal Medicine Pg no. 3363.3364,3358

102
Immunochemistry
Immunoglobulins
Immunoglobulin structure
Made up of 2 Heavy (H) and
: 2 Light (L) chains.
Disulfide bridges are present.
H chain - 440 Amino Acids(AA)
: L chain - 214 AA
5 Major classes depending on Heavy chain :
:
i. Ig G - Gamma Heavy chain
ii. Ig M - Mu Heavy chain
iii. Ig A - Alpha Heavy chain
iv. Ig D - Delta Heavy chain
v. Ig E - Epsilon Heavy chain.
V : Variable region C : Constant region

Action of proteolytic enzymes

Papain
Enzyme from Papaya
Cleaves at Hinge region and 3 fragments
:
are formed.
i. 2 Fab fragments - Antigen binding part
ii. 1 Fc fragment - Complement binding
Pepsin
Pepsin cleaves Ig at different site.
Fragments formed are F(ab)2 and Fc
: Only 2 fragments are formed.
103
Different classes of Immunoglobulins
Immunoglobulin g •
Contains 2 Heavy and 2 Light chains
•
Referred to as 7S Ig
•
Constitutes 75-80% of Total Ig
•
Antibody seen in 2 immune response
•
Can pass from vascular compartment to interstitial space.
•
Only Ig than can cross Placental barrier
•
Rh immunisation occurs due to Ig G
•
Lowest Carbohydrate content among the 5 Ig ( 2-3 %)
•
Highest T1/2 among the 5 Ig ( around 20 days )
Immunoglobulin m
•
a.k.a Macrogolubins / 19S Ig
•
5 subunits - each with 4 peptide chains
•
Joined together by J chain polypeptide
•
Very effective in agglutinating bacteria
•
Cannot come out of vascular compartment
•
Maximum distribution in vascular compartment (95%)
•
Antibodies in Primary response ( Natural antibodies )
•
Heaviest among 5 Ig
Immunoglobulin a
a.k.a 11S Ig
: Exist as Dimers usually
J chain connects dimers
: Secretory antibodies seen in seromucous secretions of
i. GI tract iv. Tears
ii. Secretory tract v. Saliva
iii. Urogenital tract vi. Sweat
Secretory piece is produced in liver and combines with
: Ig A dimer in intestinal mucosa to form secretory Ig A
Lowest Ig distributed in intra-vascular space
•
There are 2 additional subunit - S and J
Reference : Textbook of Biochemistry for Medical Students by DM Vasudevan 7th Ed
104
Immunoglobulin e
Cytophilic antibodies , 8S Ig
: 5 Heavy chain domains are present.
Mediate allergy, Hypersentivity and Anaphylaxis
: Fix on Mast cells and Basophils
Release of Histamine and Slow reacting substance leads
: to vasodilatation,hypotension and bronchoconstriction.
Peak of reaction 30 min - Immediate hypersensitivity
Immunoglobulin D
Comprise less than 1% of Serum Ig
:
7S monomer
T 1/2 - 2-3 days
: 1 variable region VH and
3 domain constant region - CH 1,CH 2,CH 3.
Very susceptible to the action of proteolytic
: enzymes at its hinge region.
Present on surface of B lymphocytes - recognition
receptors for antigens.
Reference:Textbook of Microbiology and Immunology by Subhash Chandra Parija 2nd Edition Pg :99
105
Vitamins and Minerals

Vitamin a
Interconversion of Vitamin A
Vitamin A deficiency causes -

Bitot’s spots : Grayish white triangular plaques
adherent to conjunctiva.
•
Other manifestations in eye include : Bitot’s spot

i. Night blindness or Nyctalopia
ii. Xerophthalmia
iii. Keratomalacia
•
Skin manifestation :
i. Phrynoderma
Phrynoderma
Hypercarotenemia leading to yellow

skin but no staining of sclera. Keratomalacia
106
Vitamin d
o
a.k.a Cholecalciferol
o
Sunshine vitamin
o
Anti Rachitic vitamin
o
UV - B ( 290 -315 nm )
o
Formed from pro-vitamin
“ SeCOSTEROL “
o
7 dehydrocholesterol ( an intermediate
minor pathway of cholesterol synthesis
is available in Malpighian layer of
epidermis )
AIIMS ‘20
Order in which organs

involved in Vit D synthesis :
o
Deficiency of Vitamin D
i. Rickets in children
ii. Osteomalacia in adults
Vitamin D synthesis
Rickets manifestations
Knock knee
Harrison’s Sulcus
Transverse depression passing outwards
Rickety rosary from the coastal cartilage to axilla.
Enlargement of the epiphysis at the This is due to the indentation of lower
lower end of ribs and costochondral ribs at the site of the attachment of
junction leads to beading of ribs. diaphragm Bow legs
107
Vitamin k
o
Naphthaquinone derivatives
o
Vitamin K 1 - Phylloquinone
o
Required for post translational modification of ( PLant derivative )
Coagulation factors such as
o
Vitamin K 2 - Menaquinone
i. Factor II ( Prothrombin ) (Gut flora)
ii. Factor VII ( SPCA) o
Vitamin K 3 - Menadione
iii. Factor IX ( Christmas factor ) ( Synthetic Vitamin K )
iv. Factor X ( Stuart Prower Factor )
o
Required for Gamma Carboxylation of Glutamic acid
AIIMS ‘18
Vitamin K as Co factor in GCG synthesis

o
Deficiency occurs in
i. Conditions of Malabsorption of Lipids
ii. Prolonged antibiotic therapy NEET ‘ 18 Vitamin K cycle
Quinone is regenerated by enzyme :
Dicoumarol inhibit Vit K by :
# Warfarin and Dicoumarol competitively inhibit Vitamin K

Bruise
Bruise Bleeding
# Fat Soluble vitamins - Vit. A , D , E , K

108
Vitamin b
Vitamin b 1
@
a.k.a Thiamine AIIMS ‘17
@
Physiological role
Co-enzymes for
AIIMS ‘17
i. Pyruvate dehydrogenase
AIIMS ‘17
ii. Alpha Ketoglutarate dehydrogenase
iii. Transketolase
Deficiency Manifestations
Wernicke Korsokoff Beri beri Polyneuritis

syndrome
Wet Beri Beri Dry Beri Beri Infantile

@
Cardiovascular @
CNS manifestations
manifestations
AIIMS ‘19
# Erythrocytes transketolase activity is reduced.

( Earliest manifestation seen even before clinical disturbances)
Dry Beri Beri

TPP co-enzyme for :
Wet Beri Beri
Pitting Edema Distended neck veins

Paralysis
109
Vitamin b 2
a.k.a Riboflavin
so
Heat stable
so
Flavoproteins - enzymes containing Riboflavin

so
2 co-enzymes :
so
i. FMN
ii. FAD Riboflavin structure
so
FMN Dependent enzymes :

i. Amino acid oxidation
ii. Respiratory chain
⑥
FAD Dependent enzymes :

i. Succinate dehydrogenase
ii. Acyl CoA dehydrogenase
iii. Xanthine oxidase
iv. Pyruvate dehydrogenase
v. Alpha ketoglutarate dehydrogenase
own
Riboflavin is Co factor for Glutathione
AIIMS ‘18 Co-enzymes
reductase enzyme.
# Riboflavin deficiency usually occurs other deficiency diseases such as BeriBeri,Pellagra,Kwarshiorkar.
Deficiency Manifestations of Vit B2
Proliferation of bulbar
conjunctival vessels. Glossitits, Fissured tongue and
Earliest sign of deficiency. Magenta coloured Tongue Angular stomatitis and Cheilosis
Reduced Glutathione reductase activity due to:

110
Vitamin b 3
a.k.a Niacin, Nicotinic acid , ☐
NAD + dependent enzymes

:
Pellagra preventing Factor.
Niacinamide - active form
i. LDH
ii. Glyeraldehyde-3-phosphate dehydrogenase
Co-enzyme forms : iii. Pyruvate dehydrogenase
:
i. NAD + iv. Alpha ketoglutarate dehydrogenase
ii. NADPH + v. Glutamate dehydrogenase
vi. Beta hydroxyl CoA dehydrogenase
# 60mg of Tryptophan is equivalent to 1mg of Niacin
# Quinolinate phosphoribosyl transferase is Rate limiting enzyme in conversion of Niacin to NAD +
Pellagra Causes of Niacin deficiency

Caused by deficiency of Tryptophan and Niacin. i. Dietary deficiency of Tryptophan
o
Symptoms :
o
Maize as staple diet NEET ‘20
i. Dermatitis - Casal’s Necklace
NEET ‘20 AIIMS ‘19 o
Sorghum/Johar/Guinea corn as
ii. Diarrhoea staple diet
iii. Dementia- Delirium common in A/C Pellagra ii. Deficient synthesis
iii. Isoniazid
iv. Hartnup disease
v. Carcinoid sydnrome
3. A middle aged farmer grew mainly maize as

staple food. He presents to the OPD as shown in
the image. Deficiency of which vitamin is shown?
A. Niacin C. Thiamine
B. Zinc D. Pyridoxine [NEET 2021]
Casal’s Necklace
Adverse effects
i. Flushing - transient vasodilation due to histamine release.
ii. Liver damage - modest elevations in serum transaminase and unconjugated bilirubin is seen.
iii. Diabetes - can cause hyperglycemia by insulin resistance in Diabetic patient. NEET ‘ 18 NEET ‘20
iv. Niacin Maculopathy - Thickening of Macula and Retina leading to blurred vision and blindness.
(reversible)
111
Vitamin B12
Degradation and resynthesis of methionine. [Note: The resynthesis of methionine from homocysteine is the only
reaction in which tetrahydrofolate both carries and donates a methyl (−CH3) group. In all other reactions, SAM is
the methyl group carrier and donor.] PPi = pyrophosphate; Pi = inorganic phosphate; NH3 = ammonia.
Causes of B12 Deficiency

Nutritional vitamin B12 deficiency is very common among vegetarians of low
:
socioeconomic group.
Decrease in Absorption
- Absorptive surface is reduced by gastrectomy, resection of ileum and
malabsorption syndromes.
Addisonian Pernicious Anemia
Gastric Atrophy
Fish Tape worm infection ( Diphillobothrium latum )
- This tapeworm has a special affinity to B12 causing reduction in available vitamin.
112
Deficiency Manifestations
i. Manifestations of folate deficiency
ii.Megaloblastic anemia
iii. Abnormal homocysteine level
iv.Demyelination
v.Subacute combined degeneration :
- There is demyelination affecting cerebral cortex as well as dorsal column and
pyramidal tract of spinal cord.
- Both sensory and motor tracts are affected
- Symmetrical paraesthesia of extremities, alterations of tendon and deep senses
and reflexes,loss of position sense, unsteadiness in gait, positive Romberg’s
sign (falling when eyes are closed) and positive Babinski’s sign (extensor plantar
reflex) are seen.
vi.Achlorhydria
Assessment of B12 Deficiency
i. Serum B12: It is quantitated by radioimmunoassay or by ELISA.
ii.Schilling test
iii.Methyl malonic acid: It is seen in urine.
iv.FIGLU excretion test
v. Peripheral smear: Peripheral blood and bone marrow morphology shows
megaloblastic anemia
1.Deficiency of which among the following Vitamin leads to the deficiency of THF ?
A.Vit B6 C.Vit B1
B.Vit B12 D.Vit B2 [INICET 2021]
•
Term “vitamin B12” is used as a generic descriptor for the cobalamins —those corrinoids
(cobalt-containing compounds possessing the corrin ring) having the biological activity of
the vitamin.F
Found only in foods of animal origin, there being no plant sources of this vitamin.
÷ Strict vegetarians (vegans) are at risk of developing B12 deficiency.

Normal daily requirement : 1–2 ug/day.
During pregnancy and lactation : 2 mg/day.
113
Copper
Menke’s kinky hair syndrome NEET ‘19
X Linked Recessive
•
Defective Lysyl oxidase

•
Defect in transportation of absorbed Copper

0
from from GI tract into blood.

Mutation in ATP7A gene
•
Absence of Copper binding ATPase

0
Hypothermia
•
Hair is short,fine,
•
Hypotonia
•
brittle,light coloured.
Poor Feeding
•
Hypopigmented and thin cheeks
•
Seizures
•
Depressed nasal bridge.
•
Wilson disease
•
Autosomal Recessive disorder
•
Abnormal gene is localised to long arm of Chr.13
•
Absence or Malfunction of ATP 7B
Neurologic Psychiatric
Intention tremor
•
Depression •
Dysarthria
•
Personality changes
•
Rigid dystonia
•
Anxiety •
Kayser Fleischer Ring : deposition

Parkinsonism
•
Psychosis •
of Copper in cornea
Choreiform movements
•
Diagnosis
i. Decreased Ceruloplasmin levels ( <20 mg/dL) Treatment
ii. Urinary levels of 3-Methyl Histdine is Low NEET ‘19 i. Restrict Dietary Copper intake to < 1mg/day
iii. Urinary Copper excretion is increased ii. D-Penicillamine Chelation therapy
iv. Serum Free Copper may be elevated iii. Zinc used as Adjuvant therapy
v. Liver Biopsy - hepatic copper > 250ug/g dry weight. iv. Antioxidants - Vit E
114
Heme Synthesis and Breakdown

Structure of heme
Derivative of the porphyrin.
: Porphyrins are cyclic compounds
formed by fusion of 4 pyrrole
rings linked by methenyl (=CH-)
bridges.
Reference : Textbook of Biochemistry for Medical Students by DM Vasudevan 7th Ed

BIOSYNTHESIS OF HEME
First Step in heme synthesis

ALA synthase is located in the mitochondria and is the
:
rate-limiting enzyme of the pathway.
Requires pyridoxal phosphate
Regulation of Heme Synthesis
Heme inhibits the synthesis of ALA synthase by
acting as a co-repressor.
ALA synthase is also allosterically inhibited by
:
hematin.
Ferrochelatase and ALA dehydratase are inhibited
by lead.
High cellular concentration of glucose prevents
0
induction of ALA synthase.
Reference : Textbook of Biochemistry for Medical Students by DM Vasudevan 7th Ed Pg 270

115
Shunt bilirubin
When 15N or 14C labeled glycine is injected,
: this is incorporated into heme and into

RBCs.
RBCs are lysed after 100-120 days, the
radiolabeled Hb level is decreased, along with
consequent rise in radioactive bilirubin.
However,about 15% of radioactive bilirubin is
: excreted within about 10 days.
This is called Shunt bilirubin.
Disorders of Heme Synthesis
EGG
Porphyria variegata is due to deficiency of Protoporphyrinigen oxidase.
Absorption Bands of Porphyrins

•
The 4 pyrrole rings are joined by methenyl
(-CH=) bridges and they are colored
compounds.
The double bonds are resonating and
: therefore keep shifting their position.
Heme - ferrous iron (Fe++) - red in color.
Hematin - ferric (Fe+++) - dark brown in
: colour.
All porphyrins will have an absorption band
near 400 nm; this distinguishing band is
called the Soret band.
116
CATABOLISM OF HEME
Biliverdin which is green in color.
: Bilirubin, a red-yellow pigment
Heme is degraded primarily by a microsomal
: enzyme system; heme oxygenase.
It requires molecular oxygen and NADPH.
Van Den Bergh Test for Bilirubin

Bilirubin reacts with diazo reagent (diazotized sulfanilic acid) to produce colored azo pigment.
: At pH 5, the pigment is purple in color.
Direct Reaction : Conjugated bilirubin, being water soluble gives the color immediately.
: Indirect Reaction : Free bilirubin is water insoluble. It has to be extracted first with alcohol, when the
reaction becomes positive.
Tests for Bile Pigments

Fouchet’s Test
Principle: Barium chloride reacts with sulfate radicals
:
in urine to form barium sulfate. If bilirubin is present in
urine, it adheres to the precipitate and is detected by
oxidation of bilirubin (yellow) to biliverdin (green) with
ferric chloride in the presence of trichloroacetic acid
present in Fouchets reagent.
Freshly passed urine is required.

and Dr.Aparna Verma
117
Tests for Bile Pigments and interpretation
Hays test
•
Principle:-Bile salts lower the surface tension of
urine and allow the sulfur particles
to sink.
•
Procedure: Fill 2/3rd of the test tube with urine
and sprinkle a little sulfur powder over the urine.
Note if sulfur particles sink which indicates the
presence of bile salts. Repeat with water as
control.

and Dr.Aparna Verma
118
Electron Transport Chain

Components and sequence of reactions of electron transport chain
Complex IV or Cytochrome Oxidase

i.It contains different proteins, including cytochrome a and cytochrome a3.
- The Complex IV is tightly bound to the mitochondrial membrane.
ii.Four electrons are accepted from cytochrome c, and passed on to molecular oxygen.
4 H+ + O2 + 4 Cyt. C-Fe++ → 2 H2O + 4 Cyt.C +++
iii. 2 protons are pumped out to the inter-membrane space.
iv. Cytochrome oxidase has 4 redox centers, namely, a, a3, CuA and CuB.
- Complex III or cytochrome reductase (cytochrome b-c1 complex) of respiratory
chain
Cyto c →CuA →Cyto a →Cyto a3 →CuB
•
Cytochrome oxidase contains two heme groups and two copper ions.
- The two heme groups are denoted as cytochrome-a and cytochrome a-3.
- The functional unit of the enzyme is a single protein, and is referred to as
cytochrome a--a3.
Complex IV (cytochrome oxidase) of respiratory chain

119
Inhibitors of electron transport chain and oxidative phosphorylation.
7.All of the following inhibit cytochrome oxidase except ?

A.Hydrogen sulfide C.Carbon monoxide
B.Cyanide D.Methane INICET 2021]
In an experiment conducted by a student on ETC , when pyruvate and one inhibitor is added, it
resulted in inhibition of oxidative phosphorylation. Which inhibitor most likely used by the student.
A. Oligomycin C. Antimycin
B. 2,4-DNP D. Rotinonc [NEET 2021]
Compounds which affect electron transport chain and

oxidative phosphorylation
120
121

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Biochemistry

9. Cholesterol and Lipoproteins 77

10. Lipid Storage Disorders 82

11. Molecular Genetics - Structure of DNA 85

12. DNA Replication 87

13. Different Classes of RNA 92

15. Recombinant DNA Technology 94

16. Amplification and Hybridisation Techniques 98

17. Pattern of Inheritance 101

18. Immunochemistry 102

19. Vitamins and Minerals 105

20. Heme Synthesis and Breakdown 114

21. Electron Transport Chain 118

Sub cellular organelles

Reference : Textbook of Biochemistry for Medical Students by DM Vasudevan 7th Ed. Pg 11

Fluid mosaic model

In Acidic medium they exist in Cationic.

:From the graph of Glycine,its evident that there are 2 pK values

Titration curve Histidine Titration curve Glycine AIIMS ‘19

Simple amino acid

Glycine ( Gly) Alanine ( Ala)

Branched amino acids

Valine ( Val ) Leucine ( Leu ) Isoleucine ( Ile)

Serine ( Ser) Threonine ( Thr ) Cysteine ( Cys ) Methionine ( Met )

Amino acids with amino group Dicarboxylic amino acids

Glutamine ( Gln ) Aspartic acid ( Asp ) Glutamic Acid ( Glu)

Dibasic amino acids Aromatic amino acids

Arginine ( Arg ) contains Lysine ( Lys )

Indole group Imidazole group

some Derived amino acids

Gamma carboxy Ornithine

Carbamoyl phosphate Ornithine

Trichorrhexis nodosa Nitrogen Scavengers used in Urea Cycle Disorder

Metanephrines ( Total and Fractionated )

PHENYLKETONURIA AIIMS ‘18

Caused by Phenylalanine hydroxylase deficiency

Hypopigmented hair Convulsions

Tandem Mass Spectrometry

Ferric Chloride test

Diet containing low Phenylalanine

Tyrosine levels - Normal.

Benedict’s test - Strongly Positive.

Ferric Chloride test positive.

Nitisinone combined with Phenylalanine and Tyrosine

Blackening of Urine restricted diet.

Oculocutaneous Albinism OCULAR Albinism

Sulphur content in Cystine makes crystals radio-opaque.

Cyanide Nitroprusside test showing Alkalanisation of urine by Sodium bicarbonate

Picture of carcinoid ( at center) Diagnosis

and arginine normal

Blue diaper syndrome

Reference : Nelson Testbook of Pediatrics 20th Ed. Pg 1847

Convulsions Neuroimaging - Cerebral edema

Genetic counselling,Carrier testing and

Axial MRI showing extensive Prenatal diagnosis - methods of prevention.

between residues which are about 3-4 Amino

Beta bends formed are stabilised by Intrachain

form Alpha helix.

Reference : Textbook of Biochemistry for Medical Students by DM Vasudevan 7th Ed. Pg 34

# Myoglobin - Alpha helix and Bet a pleated sheet.

TANDEM MASS SPECTROMETRY

Used to analyse complex peptide mixtures without

9.Which is the most accepted method for HbA1c estimation ?

: The column matrix is a cross-linked polymer with