PROTEIN CHEMISTRY

Dr. B.Divya Dharshini

Department of Biochemistry
 Objectives

SESSION I

• How does the structure of amino acids affect the proteins

that contain them ?

• In addition to their role as protein building blocks,

what other functions do amino acids have ?

• What are the properties of amino acids ?

• The term PROTEIN is derived from the Greek
word proteios which means primary or holding
first place.

• Major structural and functional aspects of body are

are carried out by proteins
• Proteins are macromolecules composed of many
amino acids.

• In other words proteins are polymers of amino acids.

• The sequence of the amino acids determine each

proteins unique 3 dimensional structure and its
specific function.
The Central dogma

Genes containing
codons for aminoacids

transcription

mRNA containing codon sequence

of amino acid

translation

Protein with specific amino acid sequence

(Made up of L – amino acids)
 What are amino acids ?
• Amino acids are monomers of proteins
• 300 amino acid occurs in nature
• A subset of only 20 constitute the monomeric units of a
protein
Amino acids

Common amino acids Derived amino acids

Has at least one codon Common amino acids gets

in the genetic code altered after translation process.
Eg. Alanine, glycine, cysteine Cystine, hydroxylysine,
hydroxyproline
 Biomedical importance of amino acids

In health

• Diet contains essential amino acids

• In the form of proteins amino acids perform a multitude of

structural, hormonal, catalytic functions.

• Participates in intracellular functions – nerve transmission,

regulation of cell growth, biosynthesis of porphyrins,
purines, pyrimidines, urea, creatinine, neurotransmitters
 Biomedical importance of amino acids

In disease

• Genetic defects in metabolism of amino acids results in Inborn

errors (Phenyl ketonuria)

• Additional genetic disease can result from an impaired ability

to transport specific amino acids – leads to increased excretion
of amino acids – aminoaciduria

• Inborn error of urea cycle

General Amino Acid Structure

H2N Cα COOH

R Aliphatic chain
Classification of amino acids

• BASED ON STRUCTURE

• BASED ON SIDE CHAIN CHARACTERS

• BASED ON METABOLIC FATE

• BASED ON NUTRITIONAL REQUIREMENTS

 Classification of amino acids
Based on structure

1.ALIPHATIC AMINO ACIDS

a)Mono amino mono carboxylic acids

• Simple amino acids – glycine, alanine

• Branched chain amino acids – valine, leucine, isoleucine
• Hydroxy amino acids – serine, threonine
• Sulphur containing amino acids – cysteine, methionine
•Amino acids with amide groups – asparagine
glutamine
 Structure of amino acids
Amino acids with aliphatic side groups

Glycine
(Gly / G)

Alanine
(Ala / A)

Valine
(Val / A)
 Classification of amino acids
Based on structure

1.ALIPHATIC AMINO ACIDS

a)Mono amino mono carboxylic acids

• Simple amino acids – glycine, alanine

• Branched chain amino acids – valine, leucine, isoleucine
• Hydroxy amino acids – serine, threonine
• Sulphur containing amino acids – cysteine, methionine
•Amino acids with amide groups – asparagine
glutamine
 Structure of amino acids
Amino acids with aliphatic side groups

Leucine
(Leu /L)

Isoleucine
(IIe / I)
 Classification of amino acids
Based on structure

1.ALIPHATIC AMINO ACIDS

a)Mono amino mono carboxylic acids

• Simple amino acids – glycine, alanine

• Branched chain amino acids – valine, leucine, isoleucine
• Hydroxy amino acids – serine, threonine
• Sulphur containing amino acids – cysteine, methionine
•Amino acids with amide groups – asparagine
glutamine
 Structure of amino acids

Aliphatic amino acids with hydroxyl R groups

Serine
(Ser / S)

Threonine
(Thr / T)
 Classification of amino acids
Based on structure

1.ALIPHATIC AMINO ACIDS

a)Mono amino mono carboxylic acids

• Simple amino acids – glycine, alanine

• Branched chain amino acids – valine, leucine, isoleucine
• Hydroxy amino acids – serine, threonine
• Sulphur containing amino acids – cysteine, methionine
•Amino acids with amide groups – asparagine
glutamine
 Structure of amino acids
Aliphatic amino acids with sulfur containing R group

Cysteine
(Cys / C)

Methionine
(Met / M)
 Classification of amino acids
Based on structure

1.ALIPHATIC AMINO ACIDS

a)Mono amino mono carboxylic acids

• Simple amino acids – glycine, alanine

• Branched chain amino acids – valine, leucine, isoleucine
• Hydroxy amino acids – serine, threonine
• Sulphur containing amino acids – cysteine, methionine
•Amino acids with amide groups – asparagine
glutamine
b) Mono amino dicarboxylic acids – Aspartic acid , glutamic acid

Asparatic acid &

asparagine
( Asp / D)
(Asn / N)

Glutamic
acid &
Glutamine
(Glu / E)
(Gln / Q)
Dibasic mono carboxylic acids – Lysine, arginine, histidine

Arginine
(Arg / R)

Lysine
(Lys /K)

Histidine
(His / H)
 Classification of amino acids
Based on structure

2. AROMATIC AMINO ACIDS

Phenylalanine, tyrosine
Phenylalanine
benzene (Phe / F)

phenol Tyrosine
(Tyr / Y)
 Classification of amino acids
Based on structure
3. Heterocyclic amino acids
Tryptophan, histidine

indole imidazole

Tryptophan Histidine
(Trp / W) (His/ H)

acids Proline
Proline (Pro / P)
pyrrolidine
 Classification of amino acids
Based on structure

4) DERIVED AMINO ACIDS

• Amino acids found in proteins

Hydroxy proline, hydroxy lysine

• Amino acids not seen in proteins

Citrulline, ornithine, homocysteine

• Non alpha amino acids

GABA -  amino butyric acid,  alanine
 Classification of amino acids
Based on side chain characters
1) NON POLAR SIDE CHAINS
alanine, valine, leucine, isoleucine methionine, proline,
phenylalanine, tryptophan

2) NON IONIC POLAR SIDE CHAINS

glycine, serine, threonine, Cysteine, tyrosine, glutamine,
asparagine

3) IONIC POLAR SIDE CHAINS

Acidic amino acids - Aspartic acid, glutamic acid
Basic amino acids - Lysine, arginine, histidine
Structure of amino acids

Leucine
(Leu /L)

Isoleucine
(IIe / I)
 Classification of amino acids
Based on side chain characters
1) Non polar side chains
alanine, valine, leucine, isoleucine methionine, proline,
phenylalanine, tryptophan

2) Non ionic polar side chains

glycine, serine, threonine, Cysteine, tyrosine, glutamine,
asparagine

3) Ionic polar side chains

Acidic amino acids - Aspartic acid, glutamic acid
Basic amino acids - Lysine, arginine, histidine
 Structure of amino acids
Aliphatic amino acids with hydroxyl R groups

Serine
(Ser / S)

Threonine
(Thr / T)
 Classification of amino acids
Based on side chain characters
1) Non polar side chains
alanine, valine, leucine, isoleucine methionine, proline,
phenylalanine, tryptophan

2) Non ionic polar side chains

glycine, serine, threonine, Cysteine, tyrosine, glutamine,
asparagine

3) Ionic polar side chains

Acidic amino acids - Aspartic acid, glutamic acid
Basic amino acids - Lysine, arginine, histidine
Asparatic acid
( Asp / D)

Glutamic acid
(Glu / E)

Arginine
(Arg / R)

Lysine
(Lys /K)
 Classification of amino acids
Based on metabolic fate

1) PURELY KETOGENIC
leucine

2) KETOGENIC AND GLUCOGENIC

lysine, isoleucine & aromatic amino acids

3) PURELY GLUCOGENIC
alanine, arginine, aspartic acid, cysteine, glutamic acid,
glycine, histidine, hydroxyproline, methionine, proline, serine,
threonine, valine
 Classification of amino acids
Based on nutritional significance

1) ESSENTIAL AMINO ACIDS (EAA)

- carbon skeleton cannot be synthesized in our body

- preformed AA are to be taken for normal growth
- 10 EAA
- Arginine, histidine, isoleucine, leucine, threonine, lysine,
Methionine, phenylalanine, tryptophan, valine.

Any Help In Learning These Little Molecules Proves Truly

Valuable
2) Semi essential amino acids
- partially synthesized
- growing children require them but not adults
- Histidine, arginine

3) Non-essential amino acids

- carbon skeleton can be synthesized by our body
- Alanine, aspartic acid, cysteine, glutamic acid, glycine,
proline, serine, asparagine, glutamine, tyrosine

* Selenocysteine – 21st aminoacid

* Pyrrolysine - 22nd aminoacid
 Properties of amino acids
1. PHYSICAL PROPERTIES
• Solubility – colourless, crystalline
- highly soluble in water but slightly soluble in alcohol and
insoluble in ether.
- Soluble in dilute acids and bases
• Melting point
- Very high, above 2000 C
• Sweet in taste – glycine, alanine, valine, serine,tryptophan,
histidine & proline
• Leucine is tasteless, Isoleucine & arginine are bitter
• Sodium glutamate – flavouring agent
• Aspartame (aspartic acid & phenylalanine) – artificial
sweetner
 Properties of amino acids
• Optical properties
- due to presence of asymmetric carbon/ chiral carbon
- All amino acids except glycine exhibit optical activity
because of the asymmetry at the  carbon atom.

•There are two possible arrangements for

molecules with chiral carbons.
• Molecules that differ only in spatial
arrangement of their atoms are called
stereoisomers / optical isomers / enantiomers
• The two forms are mirror images of each
other.
• Most asymmetric molecules found in
living organism occur in only one stereoisomer
form either D or L.
• All amino acids in proteins are of L-type.
• D- amino acids are found in some antibiotics
and bacterial cell wall
 Amino acids

• At physiological pH (7.4) the carboxyl group is dissociated,

forming the negatively charged carboxylate ion.

• The amino group is protonated.

• In proteins, almost all of these carboxyl and amino groups

are combined in peptide linkages and are not available for
chemical reactions (Except for hydrogen bonding)

• Thus the nature of the side chains is that which

utlimately dictates the role an amino acid plays in a protein.
Properties of amino acid –Ampholyte and Isoelectric pH

•AA are Amphoteric in nature

•Exist as Ampholytes or Zwitter ion

NH2 (basic group or proton acceptor)

H - C – COOH (acidic group or proton donor)

NH3+

H C COO –

H
• pH at which molecule carries no net charge is known as
isoelectric point or isoelectric pH
• Concentration of zwitterion is maximum at this pH
• Acidic solutions- cationic form
• Alkaline- anions
• Amino acid , peptides, proteins has a particular pI
• Knowledge of pI helps in separation of amino acids or
peptides or proteins

• ISOELECTRIC pH(PI)
- no net charge
- all groups ionised and charges cancel each other
- no mobility in electric field
- minimal solubility and buffering capacity
 Ionic property

K1 K2
+
H3N-CH3-COOH H+ + + H3N-CH2-COO- H+ + H2N-CH2-COO-

CATION ZWITTERION ANION

pI – isoeletric pH ( which is usually the mean of all the pK values for that amino acids)

pI = pK1 + pK2
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2
 General reactions of amino acids
1. CHEMICAL PROPERTIES

a) Reactions due to carboxyl group

* ALPHA DECARBOXYLATION TO FORM AMINES – removal of

CO2
• Histidine – Histamine
• Tyrosine - Tyramine
• Tryptophan – Tryptamine
• Lysine - Cadaverine
 General reactions of amino acids
* AMIDE FORMATION
- Dicarboxylic amino acid forms amides with ammonia
• Aspartic acid – asparagine
• Glutamic acid - Glutamine
b) Reactions due amino group

* TRANSAMINATION
• transfer of  amino group to  keto acid to form new amino acid
and keto acid
• interconversion of amino acids & synthesis of non essential
amino acids
* OXIDATIVE DEAMINATION
• removal of  amino group from AA as ammonia and keto acid formed
– glutamic acid

* CARBAMINO COMPOUND
• CO2 added to alpha amino group of AA to form carbamino compound
– transport of CO2 from tissues to the lungs by haemoglobin

Hb-NH2 + CO2 Hb-NH-COOH (carbamino Hb)

c) Reactions due to side chains

* TRANSMETHYLATION
The methyl group of methionine is used for transmethylation
reactions.

* ESTER FORMATION
Ester formation by OH group –serine and threonine forms the
phosphorylated form (phospholipids). It also involved in the
synthesis of glycoproteins.
* REACTIONS OF AMIDE GROUP
Glutamine and asparagine forms N-glycosidic bonds with
carbohydrates to form glycoproteins.

* REACTIONS OF SH GROUP
- cysteine has SH group and forms disulphide bonds (S-S)
with another cysteine
- Inter and intra disulphide bonds in the polypeptides can
be formed by two cysteine residues.
Formation of disulphide bridges
A CHAIN

SH SH

SH SH

B CHAIN

A CHAIN

S S

S S
B CHAIN

Inter disulphide linkage

 Colour reactions of amino acids

• Biuret reaction – Tripeptides & more

• Ninhydrin – α- amino acids
• Xanthoproteic – Aromatic amino acids (benzene )
• Millon’s - tyrosine (phenol)
• Aldehyde - Tryptophan (indole)
• Sulphur -Sulphur containing amino acids
• Sakaguchi – Arginine (guanidino)
• Pauly’s – Histidine (imidazole)
Biologically important products derived aminoacids

AMINOACID DERIVATIVE FUNCTION

Glutamic acid GABA Neurotransmitter
Tyrosine Dopamine Neurotransmitter
Glutamic acid Gabapentin Precursor of GABA
Histidine histamine Mediator for
allergic reaction
Tyrosine Thyroxine Thyroid hormone
Serine Cycloserine Anti TB drug
Serine Azaserine Anticancer drug
Arginine Ornithine Urea synthesis
Citrulline
 Biologically important peptides

Name Amino acid Source & Function

residues
Thyrotropin 3 Secreted by hypothalamaus, causes anterior pituitary
releasing to release thyrotropic hormone
hormone
Vasopressin 9 Secreted by posterior pituitary, causes kidney to
retain water from urine
Enkephalin 5 Opiate-like peptide found in brain that inhibits sense
of pain.
Angiotensin I, 10 Hypertensive peptide – stimulates the release of
II 8 aldosterone from adrenal cortex

Glutathione 3 Metabolism of xenobiotics

 Biologically important peptides

Name Amino acid Source and function

residues

Bradykinin 9 Vasodilator

Substance P 10 Neurotransmitter

Tyrocidin, Cyclic Polypeptides antibiotics

gramicidin peptides (10)

Aspartame 2 Sweetening agent in many beveragers

 Separation technique
CHROMATOGRAPHY
The term is derived from the Greek word “Chroma” meaning
colour.
This method was first employed by Tswett in 1903.

Paper chromatography & Thin layer chromatography

Both these are Liquid – Liquid partition chromatography.
Paper Chromatography is widely used to separate amino acids
and sugars.
Thin layer chromatography is used to separate lipids.
 Separation techniques
Paper chromatography
Types – Ascending & Descending
Stationary phase – water held on a solid support.
Mobile phase – Butanol –acetic acid –water
Principle of separation
The difference in distribution of the amino acids / sugars between two phases.
Visualisation of chromatography – after the chromatographic run the
paper is dried and it is sprayed with ninhydrin for amino acids, sulphuric
acid for phospholipids and diphenylamine for sugars
Importance of Rf - It helps in identification of the unknown
Rf = Distance traveled by solute
------------------------------------
Distance traveled by solvent
Paper chromatography
Relative Front (Rf)
Thank you