Medical Biochemistry

Yussif Adams (PhD)

Enzymes

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 Catalyst

• substance that increase rates of a

chemical reaction
• does not effect equilibrium
• remain unchanged in overall
process
• reactants bind to catalyst,
products are released

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Catalysts increase product formation by
(1) lowering the energy barrier (activation energy) for the product to form
(2) increases the favorable orientation of colliding reactant molecules for
product formation to be successful (stabilize transition state intermediate)

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 Catalytic Power

• Enzymes can accelerate reactions as much

as 1016 over uncatalyzed rates!
• Urease is a good example:
– Catalyzed rate: 3x104/sec
– Uncatalyzed rate: 3x10 -10/sec
– Ratio is 1x1014 !

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 Specificity
• Enzymes selectively recognize proper
substrates over other molecules
• Enzymes produce products in very high
yields - often much greater than 95%
• Specificity is controlled by structure - the
unique fit of substrate with enzyme
controls the selectivity for substrate and
the product yield

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 Classes of enzymes
1. Oxidoreductases = catalyze oxidation-reduction reactions
(NADH)
2. Transferases = catalyze transfer of functional groups
from one molecule to another.
3. Hydrolases = catalyze hydrolytic cleavage
4. Lyases = catalyze removal of a group from or addition of
a group to a double bond, or other cleavages involving
electron rearrangement.
5. Isomerases = catalyze intramolecular rearrangement.
6. Ligases = catalyze reactions in which two molecules are
joined.
Enzymes named for the substrates and type of reaction

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 PROSTHETIC GROUPS
• Many enzymes contain small nonprotein
molecules and metal ions that participate
directly in substrate binding or catalysis.
Termed prosthetic groups, cofactors, and
coenzymes.
• Prosthetic groups are distinguished by their
tight, stable incorporation into a protein’s
structure by covalent or noncovalent forces e.g.
pyridoxal phosphate, flavin mononucleotide
(FMN), flavin dinucleotide (FAD), thiamin
pyrophosphate, biotin, and the metal ions of Co,
Cu, Mg, Mn, Se, and Zn (metalloenzymes).
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 COFACTORS
• They bind in a transient, dissociable manner
either to the enzyme or to a substrate such as
ATP.

• Cofactors must be present in the medium

surrounding the enzyme for catalysis to occur.
• The most common cofactors also are metal ions.

• Enzymes that require a metal ion cofactor are

termed METAL-ACTIVATED ENZYMES to
distinguish them from the METALLOENZYMES
for which metal ions serve as prosthetic groups.
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 COENZYMES
• They serve as recyclable shuttles—or group
transfer elements—that transport many
substrates from their point of generation to
their point of utilization.
• Association with the coenzyme also stabilizes
substrates such as hydrogen atoms or hydride
ions.
• Other substance transported are methyl groups
(folates), acyl groups (coenzyme A), and
oligosaccharides (dolichol) – thiamin, riboflavin,
niacin, biotin
• Enzyme + Co-enzyme = holoenzyme
• Enzyme alone = apoenzyme 10
 ACTIVE SITE

• Three dimensional
• Small part of the enzyme
• Cleft or crevices
• Bound to substrate through multiple weak
interaction
• Specificity depends on spatial arrangement
of atoms within the active site

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 Enzyme Inhibition
• Inhibitor – substance that binds to an enzyme and
interferes with its activity
• Can prevent formation of ES complex or prevent ES
breakdown to E + P.
• Irreversible and Reversible Inhibitors
• Irreversible inhibitor binds to enzyme through covalent
bonds (binds irreversibly)
• Reversible Inhibitors bind through non-covalent interactions
(disassociates from enzyme)
• Why important?

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 Reversible Inhibitors

E + S <-> ES -> E + P
E + I <-> EI
Ki = [E][I]/[EI]
• Competitive
• Uncompetitive
• Non-competitive

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Types of Reversible Enzyme Inhibitors

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Introduction to Metabolism

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 Metabolism
Metabolism consists of catabolism and
anabolism
Catabolism: degradative pathways
– Usually energy-yielding!
Anabolism: biosynthetic pathways
– energy-requiring!

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17
 Catabolism and Anabolism
Catabolic pathways converge to a few
end products
Anabolic pathways diverge to
synthesize many biomolecules
Some pathways serve both in
catabolism and anabolism
Such pathways are amphibolic

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19
 Organization in Pathways
Pathways consist of sequential steps
The enzymes may be;
• separate
• Or may form a multienzyme complex
• Or may be a membrane-bound system

New research indicates that

multienzyme complexes are more
common than once thought
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 Mutienzyme complex

Separate
enzymes

Membrane
Bound System

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 Organization of Pathways
Closed Loop
(intermediates recycled)

Linear Spiral
(product of rxns are (same set of
substrates for enzymes used
subsequent rxns) repeatedly)
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 Metabolism Proceeds in Discrete
Steps
Enzyme specificity defines
biosynthetic route
Controls energy input and
output
Allow for the establishment of
control points.
Allows for interaction between
pathways

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Regulation of Metabolic Pathways
• Pathways are regulated to allow the organism to
respond to changing conditions.
• Most regulatory response occur in millisecond
time frames.
• Most metabolic pathways are irreversible under
physiological conditions.
• Regulation ensures unidirectional nature of
pathways.
• Flow of material thru a pathway is referred to
as flux.
• Flux is regulated by supply of substrates,
removal of products, and activity of enzymes

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 Enzyme Regulation of Flux
Common mechanisms
• feedback inhibition – product of pathway down
regulates activity of early step in pathway

• Feedforward activation – metabolite produced

early in pathway activates down stream enzyme

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 CARBOHYDRATE METABOLISM AND
DISORDERS
• The main function of glucose is a major tissue energy
source through the glycolytic pathway and the kreb’s
cycle.
• The brain depends on extracellular glucose concentration
for its energy therefore in situations of low glucose
levels, there is impaired cerebral function which may even
lead to an irreversible neuronal damage.
• The brain cannot:
– Synthesize glucose
– Store significant amounts of glucose
– Metabolize substrates other than glucose and ketones
– Extract enough glucose from ECF at low concentrations for its
metabolic needs
 Blood glucose homeostasis
Glucose enters the blood through
• Dietary carbohydrates
• Gluconeogenesis
• Hepatic glycogenolysis

Glucose leaves the blood through the ff:

• Its oxidation to carbon dioxide and water
• Glucose conversion to fatty acids in adipose tissues.
• Glycogenesis
• Excretion when the renal threshold is exceeded
OVERVIEW OF GLUCOSE METABOLISM & HOMOESTASIS
 1. Dietary carbohydrates
Glycolysis
• It occurs in the cytoplasm of the cell and does
not require the presence of oxygen (Anaeorbic
process)
• Converts hexose to two pyruvates
• Generates 2 ATP and 2 NADH
• For certain cells in the brain and eye, glycolysis is
the only ATP generating pathway

Glucose+2ADP+2NAD++2Pi -> 2pyruvate+2ATP+2NADH+2H++2H20

 Glycolysis
• Essentially all cells carry out glycolysis
• Ten reactions - same in all cells - but rates
differ
• Two phases:
– First phase converts glucose to two G-3-P
– Second phase produces two pyruvates
• Products are pyruvate, ATP and NADH
• Three possible fates for pyruvate

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 Rx1: Hexose Kinase
• 1st step in glycolysis; G large, negative
• This is a priming reaction - ATP is consumed here
in order to get more later
• ATP makes the phosphorylation of glucose
spontaneous
• Require Mg/Mn as cofactor

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Hexokinase also functions in other processes

Not 1st committed

step in glycolysis
Glucose import

Directing glucose to
other pathways33
 Different Hexokinase Isozymes
• Two major forms hexokinase (all cells) &
glucokinase (liver)
• Km for hexokinase is 10-6 to 10-4 M; cell has 4
X 10-3 M glucose
• Km for glucokinase is 10-2 M only turns on when
cell is rich in glucose
• Glucokinase functions when glucose levels are
high to sequester glucose in the liver.
• Hexokinase is regulated - allosterically
inhibited by (product) glucose-6-P

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 Rx 3: Phosphofructokinase
PFK is the committed step in glycolysis!
• The second priming reaction of glycolysis
• Committed step and large, -DG – means PFK is highly
regulated
• b-D-fructose-6-phosphate is substrate for rxn

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Phosphofructokinase is highly regulated
• ATP inhibits, AMP reverses inhibition
• Citrate is also an allosteric inhibitor
• Fructose-2,6-bisphosphate is allosteric
activator
• PFK increases activity when energy status is
low
• PFK decreases activity when energy status is
high

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 Rx 6: Glyceraldehyde-3P-Dehydrogenase
• G3P is oxidized and phosphorylated to 1,3-BPG
• Near equilibrium rxn
• Pi is used as phosphate donor
• C1 phosphoryl group has high group transfer potential,
used to phosphorylate ADP to ATP in next step of
glycolysis
• Arsenate can replace phosphate in rxn (results in lower
ATP)
• NADH generated in this reaction is reoxidized by
respiratory electron transport chain (generates ATP)

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 Rx 7: Phosphoglycerate Kinase (PGK)
• ATP synthesis from a high-energy phosphate
• This is referred to as "substrate-level
phosphorylation"
• Although has large negative DGo’ (-18 kJ/mole) because
PGK operates at equilibrium in vivo, the overall DG is 0.1
Kj/mole and is a near-equilibrium rxn.
• 2,3-BPG (for hemoglobin) is made by circumventing the
PGK reaction

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 Rx 9: Enolase
• Near equilibrium rxn
• "Energy content" of 2-PG and PEP are similar
• Enolase just rearranges to a form from which
more energy can be released in hydrolysis
• Requires Mg2+ for activity, one bings Carboxyl
group of substrate the other involved in catalysis.

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 Rx 10: Pyruvate Kinase
• Substrate level phosphorylation generates
second ATP
• Require Mg/Mn/K
• Large, negative G - regulation!
• Allosterically activated by AMP, F-1,6-bisP
• Allosterically inhibited by ATP, long chain
FA, citrate and acetyl-CoA

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 Pyruvate can go in three major
directions after glycolysis
• Under aerobic conditions pyruvate is oxidized to
Acetyl-CoA which can enter Citric acid (TCA)
cycle.
• Under anaerobic conditions pyruvate can be
reduced to ethanol (fermentation) or lactate
• Under anaerobic conditions formation of ethanol
and lactate is important in the oxidization NADH
back to NAD+
• Under aerobic conditions NADH is oxidized to
NAD+ by the respiratory electron transport chain.

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 Lactate formation
NADH NAD
O OH
O O
H3C C C H3C C C
H
O O
NADH NAD

•In animals under anaerobic conditions pyruvate

is converted to lactate by the enzyme lactate
dehydrogenase
•Impt for the regeneration of NAD+ under
anaerobic conditions.
 Alcohol Fermentation

•Important for the regeneration of NAD+ under

anaerobic conditions
•Process common to microorganisms like yeast
•Yields neutral end products (CO2 and ethanol)
•CO2 generated impt in baking where it makes dough
rise and brewing where it carbonates beer.
Other Sugars can
enter glycolysis
How other sugars enter glycolysis

• Mannose can be phosphorylated to mannose-6-

phosphate by hexokinase and then converted to
fructose-6-phosphate by phosphomannose
isomerase.
• Fructose can be phosphorylated by fructokinase
to form fructose-1 phosphate (F-1-P). F-1-P
can then be converted to glyceraldehyde and
DHAP by F-1-P aldolase. Triose kinase then
converts glyceraldehyde to G-3-P.
 Fructosuria

• Deficiency of fructokinase, fructose exceed

renal threshold, may be confused with glucosuria

• May give false positive test for glucose using

only urine dipstick

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 Fructose Intolerance

• Serious autosomal recessive due to

deficiency of fructose-1-P aldolase in liver
and kidney
• Accumulation of fructose-1-P inhibit the
action of fructokinase
• Infant fail to thrive, hepatosplenomegaly,
jaundice, convulsion and coma at weaning
when sucrose is added to diet
• There could also be repeated vomitting, loss
in weight, sweating and lethargy (weakness)
• Diagnosis: fructosuria and hypoglyceamia
after ingesting sucrose 48
 Galactose

• Obtained by hydrolysis of milk sugar,

lactose
• Galactokinase is highly specific for D-
galactose
• Inducible enzyme whose activity is
increase by feeding with galactose

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 Galactosemia
• Inherited as an autosomal recessive
• Deficiency of galactose-1- phosphate
uridylytransferase.
• galactose-1-phosphate accumulates
• Leads to liver damage, GIT, Kidney
and brain
• Untreated infants fail to thrive often
have mental retardation.
• Can be treated with galactose free
diet.
 Cataracts
• Accumulation of galactose in the blood
and subsequent spillage in urine and
other tissue
• In the lens of the eye, galactose is
change to a polyol (galactitol/dulcitol)
by galactose reductase
• Gal +NADPH +H → Galactitol + NADP+
• Accumulation of dulcitol in the lens
damage tissue via opacification i.e.
cataracts
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 Lactose Intolerance
• Humans undergo reduction in lactase at 5
to 7 years of age.
• In lactase deficient individuals, lactose is
metabolized by bacteria in the large
intestine.
• Produce CO2, H2 and short chain acids.
• Short chain acids cause ionic imbalance in
intestine (diarrhea)
Control Points in
Glycolysis

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Deregulation of Glycolysis in Cancer
Cells
• Glucose uptake and glycolysis is ten times faster in
solid tumors than in non-cancerous tissues.
• Tumor cells initally lack connection to blood supply
so limited oxygen supply
• Tumor cells have fewer mitochondrial, depend more
on glycolysis for ATP
• Increase levels of glycolytic enzymes in tumors
(oncogene Ras and tumor suppressor gene p53
involved)

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2. Gluconeogenesis

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 Gluconeogenesis
• Synthesis of "new glucose" from common
metabolites (pyruvate, lactate, citric acid cycle
intermediates, amino acids other than lysine or
leucine, and glycerol)
• Humans consume 160 g of glucose per day
• 75% of that is in the brain
• Body fluids contain only 20 g of glucose
• Glycogen stores yield 180-200 g of glucose
• The body must still be able to make its own
glucose

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 Gluconeogenesis
• Occurs mainly in liver and kidneys
• Not the mere reversal of glycolysis
for 2 reasons:
– Energetics must change to make
gluconeogenesis favorable
– Reciprocal regulation must turn one on
and the other off - this requires
something new!

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• Seven steps of glycolysis
are retained
• Three steps are replaced
• The new reactions
provide for a
spontaneous pathway (G
negative in the direction
of sugar synthesis), and
they provide new
mechanisms of regulation

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 Pyruvate Carboxylase
• The reaction requires ATP and bicarbonate as
substrates
• Biotin cofactor
• Acetyl-CoA is an allosteric activator
• Regulation: when ATP or acetyl-CoA are high,
pyruvate enters gluconeogenesis

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 PEP Carboxykinase
• Lots of energy needed to drive this reaction!
• Energy is provided in 2 ways:
– Decarboxylation is a favorable reaction
– GTP is hydrolyzed
• GTP used here is equivalent to an ATP

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Fructose-1,6-bisphosphatase
• Thermodynamically favorable - G
in liver is -8.6 kJ/mol
• Allosteric regulation:
– citrate stimulates
– fructose-2,6--bisphosphate inhibits
– AMP inhibits

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 Glucose-6-Phosphatase
• Presence of G-6-Pase in ER of liver and kidney cells
makes gluconeogenesis possible
• Muscle and brain do not do gluconeogenesis
• G-6-P is hydrolyzed as it passes into the ER
• ER vesicles filled with glucose diffuse to the plasma
membrane, fuse with it and open, releasing glucose
into the bloodstream.

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•Metabolites other than
pyruvate can enter
gluconeogenesis
•Lactate (Cori Cycle)
transported to liver for
gluconeogenesis
•Glycerol from
Triacylglycerol catabolism
•Pyruvate and OAA from
amino acids
(transamination rxns)
•Malate from glycoxylate
cycle -> OAA ->
gluconeogenesis
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 Regulation of Gluconeogenesis
• Reciprocal control with glycolysis
• When glycolysis is turned on, gluconeogenesis
should be turned off
• When energy status of cell is high, glycolysis
should be off and pyruvate, etc., should be
used for synthesis and storage of glucose
• When energy status is low, glucose should be
rapidly degraded to provide energy
• The regulated steps of glycolysis are the
very steps that are regulated in the reverse
direction!
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66
 Glycogenesis
(synthesis of glycogen)
 Metabolism of Tissue Glycogen
• Glycogen, a branched polymer of glucose, represents
a storage form of glucose
• Glycogen metabolism occur primarily in liver &
skeletal muscle. Cardiac muscle and the kidney, store
smaller quantities
• Glycogens stored in the cytoplasm as either single
granules (skeletal muscle) or as clusters of granules
(liver)
Glucokinase
 Glycogen Synthase
• Forms -(1 4) glycosidic bonds in glycogen

• Glycogen synthesis depends on sugar nucleotides

UDP-Glucose

• Glycogen synthase transfers glucosyl units from

UDP-glucose to C-4 hydroxyl at a nonreducing end
of a glycogen strand.
3. Hepatic glycogenolysis
 Glycogen Breakdown
• Tissue glycogen is an important energy reservoir -
its breakdown is carefully controlled
• Glycogen consists of "granules" of high MW
• Glycogen phosphorylase cleaves glucose from the
nonreducing ends of glycogen molecules
• This is a phosphorolysis, not a hydrolysis
• Metabolic advantage: product is a sugar-P - a
"sort-of" glycolysis substrate
Glucokinase
•Glycogen phosphorylase
cleaves glycogen at non-
reducing end to generate
glucose-1-phosphate
•Debranching of limit dextran
occurs in two steps.
•1st, 3 α-1,4 linked glucose
residues are transferred to
non-reducing end of glycogen
•2nd, amylo-1,6-glucosidase
cleaves 1,6 linked glucose
residue.
•Glucose-1-phosphate is
converted to glucose-6-
phosphate by
phosphoglucomutase
 Control of Glycogen Metabolism
• A highly regulated process, involving reciprocal
control of glycogen phosphorylase (GP) and
glycogen synthase (GS)
• GP allosterically activated by AMP and inhibited
by ATP, glucose-6-P and caffeine
• GS is stimulated by glucose-6-P
• Both enzymes are regulated by covalent
modification - phosphorylation
G-6-P as a metabolic junction
 Pentose Phosphate Pathway
Also known as:
• Pentose shunt
• Hexose monophosphate shunt
• Phosphogluconate pathway

• It occurs in the cytosol.

 Pentose Phosphate Pathway
• Provides NADPH for biosynthesis

• Produces ribose-5-P for RNA and DNA

• oxidative steps (formation of NADPH) followed by

non-oxidative steps

• Active in tissues that synthesis fatty acids and

sterols (liver, mammary glands, adrenal glands,
adipose tissue)

• Active in red blood cells to maintain heme in

reduced form.
Oxidative Stage

Non-oxidative
Stage
 Regulatory enzyme

5 carbon atoms
 Glutathione and NADPH
• What is glutathione?
• Why is it important?
• How is it related to NADPH?
85
 Glutathione Functions -1
• It serves as a reductant.
• Conjugates to drugs making them
water soluble.
• Involved in amino acid transport
across cell membranes.
• Cofactor in some enzymatic
reactions.
– rearrangement of protein disulfide
bonds.
 Glutathione Functions -2

• The sulfhydryl of GSH is used to

reduce peroxides (ROS) formed
during oxygen transport.
– Reactive oxygen species (ROS) damage
macromolecules (DNA, RNA, and protein)
and ultimately lead to cell death.
• The resulting oxidized form of GSH
is two molecules linked by a disulfide
bridge (GSSG).
 Glutathione and Erythrocytes
• GSH is extremely important
particularly in the highly oxidizing
environment of the red blood cell.
• Mature RBCs have no mitochondria and
are totally dependent on NADPH from
the pentose phosphate pathway to
regenerate GSH from GSSG via
glutathione reductase.
• In fact, as much as 10% of glucose
consumption, by erythrocytes, is
mediated by the pentose pathway.
Glutathione and Erythrocytes
• The reduced form of glutathione
serves as a sulfhydryl buffer.
• It maintains cysteine residues in
hemoglobin and other proteins in a
reduced state.
• GSH is essential for normal RBC
structure and keeping hemoglobin in
Fe++ state.
Glutathione and Erythrocytes
• Reduced glutathione also detoxifies
peroxides.
2GSH + ROOH  GSSG + H2O + ROH
• Cells with low levels of GSH are
susceptible hemolysis.
• Individuals with reduced GSH are
subject to hemolysis.
• This is often clinically seen as black
urine under certain conditions.
Conditions for hemolytic anemia related
G6PD deficiency.
• The ingestion of oxidative agents that
generate peroxides or reactive oxygen
species (ROS).
– Antimalarials - pamaquine
– purine glycoside from fava beans.
• Individuals with G6PD deficiency can not
produce sufficient GSH to cope with the
ROS.
• Proteins become cross linked leading to
Heinz body formation and cell lysis.
• People with the disorder are not normally anemic
and display no evidence of the disease until the
red cells are exposed to an oxidant or stress.
Drugs that can precipitate this reaction:
• antimalarial agents
• sulfonamides (antibiotic)
• aspirin
• nonsteroidal anti-inflammatory drugs (NSAIDs)
• nitrofurantoin
• quinidine
• quinine
• exposure to certain chemicals - mothballs
 FAVISM
• Individuals with G6PD deficiency must
not eat Fava beans.
• Erythrocytes lyse=dark or black urine.

• Interesting
– The growth Plasmodium falciparum (malaria
parasite) fails in G6PD deficient individuals.
Gylcolysis

Electron Transport and

Oxidative phosphorylation

TCA Cycle
 Pancreatic Anatomy and Function

• The pancreas has two major functions: production and release

of digestive enzymes, and production and release of the two
major hormones responsible for the endocrine control of
glucose metabolism namely insulin and glucagon.

• The exocrine pancreas comprises about 98% of the mass of

the organ.

• The acinar cells synthesize and secrete the digestive enzymes

(trypsin, chymotrypsin, elastase, amylase, and others) that
break down food into simpler components that can be
absorbed by the intestine.

• The endocrine pancreas is comprised of small groups of cells

distributed throughout the organ. These groups, the Islets of
Langerhans, consist of four cell types (α, β, δ, and F).
• The islet composition varies in different parts of the
pancreas (posterior primarily F-cells with some β-
cells, and anterior primarily β-cells with some α and
amounts of δ-cells).

• Each cell type secretes one major hormone: the β-cells

are the insulin producing cells, while the α-cells
produce glucagon, the δ-cells produce somatostatin,
and the F cells produce Pancreatic Polypeptide.

• The function of pancreatic somatostatin appears to be

limited to inhibition of insulin and glucagon release;
while the function of Pancreatic Polypeptide is
unknown.
 Physiological Response To Increase In Blood
Glucose
• Release of insulin by the -cells of the islets
of Langerhans
• Inhibition of the release of glucagons
• Increase uptake of glucose by the liver and
brain
• Increased uptake of glucose by the
peripheral tissues

NB: The most important hormone concerned

with glucose homeostasis is insulin
 Effect of insulin on increased blood glucose

 It increases the permeability of cell membrane

to glucose
 Insulin causes glucose transporting channels
(GLUT4) in the muscle and adipose tissues to move
from the cytosol to the cell membrane to
facilitate the uptake of glucose by the cells.

Once inside the cells, glucose becomes phosphorylated

and converted to glycogen or used for energy.

The process of phosphorylation, traps glucose and

commits it to further metabolism within the cells
 Promotes glycolysis
Insulin promotes glycolysis by stimulating formation of
a glucose metabolite, fructose 2,6-bisphosphate.

The compound is a potent activator of

phosphofructokinase-1 (PFK-1), the rate limiting
step in glycolysis.

Also, fructose 2,6-bisphosphate inhibits the activity of

1,6 biphosphatase, an enzyme which catalyzes the
reverse reaction of PFK-1.

Thus glucose metabolized to pyruvate cannot be

converted back to glucose through gluconeogenesis
 Effect of insulin on increased blood glucose

 Increase glycogenesis
In the liver, insulin does not affect glucose
transport directly (as it does in muscle and fat),
but promotes glucose uptake by stimulating its
storage as glycogen through the action of
glycogen synthase.

Glycogen synthase catalyses the synthesis of

glycogen while simultaneously deactivating the
enzyme responsible for glycogen breakdown
called glycogen phosphorylase
Glucokinase
 Effect of insulin on increased blood glucose
 Increased lipogenesis

Acetyl CoA carboxylase catalyses the formation of malonyl CoA, the

first and rate limiting step in the de novo synthesis of FA and
is activated by insulin.

Malonyl CoA inhibits the breakdown of newly formed FA by

preventing their transport into the mitochondria by inhibiting
the FA transporter protein carnitine palmitoyl transferase I .

The newly formed FA are esterified to TG and packaged as fat

rich particles called VLDL and are exported into the
bloodstream to be taken up by muscle or stored in adipose
tissues.
 Effect of counter regulatory hormones on blood
glucose
• Glucagon (29 A.A hormone secreted by -cells). This is
the first hormone to be released when glucose levels
falls hence important in the hourly maintenance of
adequate glucose levels between meals and at night. It
functions by stimulating an increase in glycogenolysis
and promotes gluconeogenesis

• Growth hormone- It is a peptide hormone from the

anterior pituitary.
It inhibits glucose uptake by the tissues and also inhibits
the synthesis of fat from carbohydrates and also
promotes the release of free fatty acids and glycerol
i.e hydrolysis TG to FFA and glycerol in adipose tissues
thereby providing an alternative fuel to glucose
• Catecholamine e.g Adrenaline/ Epinephrine:
This is a tyrosine derived molecule from the
adrenal medulla that counteracts
hypoglycaemia by:

a. Stimulating glycogenolysis in liver and

muscle cells
b. Stimulating adipocyte lipolysis by action of
hormone sensitive lipase
c. Directly inhibiting insulin release
• Thyroid hormones (T3 and T4) increase the
absorption of glucose from the GIT and stimulates
glycogenolysis. They may also accelerate the
degradation of insulin

• Glucocorticoids e.g. Cortisol

It is a steroid hormone released from the adrenal
cortex in response to ACTH.
It functions by stimulating hepatic gluconeogenesis
and inhibits glucose metabolism at the peripheral
tissues.
 KETONE BODIES [KBs]
 These are water soluble organic compounds generated in
human body under certain metabolic conditions. As water
soluble compounds, they do not require any special carrier
proteins like lipoproteins or albumin and thus are regarded
as water soluble forms of fat.

 The compounds acetone, acetoacetate and β–hydroxy

butyrate are known as ketone bodies.

 Ketone bodies are formed from acetyl CoA resulting from β

oxidation of FA in excess of optimal function of Kreb's
cycle.

 The formation of ketone bodies in the liver mitochondria is

termed ketogenesis.
 IMPORTANCE AND FUNCTIONS
• Ketone bodies being water soluble are easily transported from liver
to various tissues.

• The two KBs, acetoacetate and β–hydroxy butyrate serve as

important sources of energy for peripheral tissues like skeletal
muscle, cardiac muscle, renal cortex etc.

• The production and utilization of KBs becomes more significant when

glucose is in short supply to tissues as observed in starvation and
diabetes mellitus

• During prolonged starvation KBs are the major source of fuel for
brain and other parts of CNS. After the diet has been changed to
lower blood glucose for 3 days, the brain gets 25% of its energy
from ketone bodies. After about 40 days, this goes up to 70%, but
can not utilize FA.
 1. Formation of Citrate by
Citrate Synthase
2. Citrate to Isocitrate by
Aconitase
3. Isocitrate to alpha-
ketoglutarate by Isocitrate
Dehydrogenase
4. Alpha ketoglutarate to
Succinyl-coA by Alpha
ketoglutarate
Dehydrogenase
5. Succinyl-coA to Succinate
by Succinyl-coA synthetase
6. Succinate to Fumarate by
Succinate Dehydrogenase
7. Fumarate to Malate by
Fumarase
8. Malate to Oxaloacetate by
Malate Dehydrogenase
 Stepwise synthesis of Ketone bodies

1.Two molecules of acetyl CoA react with

each other in the presence of thiolase
enzyme to form acetoacetyl CoA.
2. Condensation of acetoacetyl CoA with
acetyl CoA to form HMG CoA (3 or β
hydroxyl- 3 or β methyl glutaryl CoA)
catalyzed by HMG CoA synthase
3. HMG-CoA lyase enzyme catalyzes the
cleavage of HMG-CoA to acetoacetate
and acetyl CoA.
4. Acetoacetate produces β-hydroxybutyrate
in a reaction catalyzed by β-
hydroxybutyrate dehydrogenase in the
presence of NADH.
5. Both acetoacetate and β-hydroxybutyrate can be
transported across the mitochondrial membrane and the
plasma membrane of the liver cells, enter to the blood
stream to be used as a fuel by other cells of the body.

6. In the blood stream, small amounts of acetoacetate are

spontaneously (non- enzymatically) decarboxyated to
acetone.
 Synthesis of Ketone bodies - ketogenesis
• Site of Ketogenesis:
• It occurs in liver and the enzymes responsible for it are
located in mitochondrial matrix
• Steps
• Formation of Acetoacetyl CoA. Enzyme involved is β-
Thiolase.
• Formation of HMG-CoA. Enzyme involved is HMG Co A
Synthase which also regulates KB synthesis.

• Breakdown of HMG-CoA to acetoacetate and acetyl CoA

by HMG-CoA lyase.

• Spontaneous breakdown of Acetoacetate to Acetone.

• Formation of β-hydroxy butyrate by enzyme β–hydroxy

butyrate dehydrogenase
 Ketoacidosis
• In normal humans there is a constant production of
ketone bodies by liver and their utilization by extra
hepatic tissues.

• The blood level of KBs is about 1 mg/dl in a normal human

adult. Their excretion in urine is very low and
undetectable by routine tests.

• When the rate of synthesis of ketone bodies exceeds

the rate of utilization, their level increases in blood. This
is known as ketonemia.

• Ketonemia is produced due to increased production

rather than decreased utilization.
• In ketonemia, the excretion of KBs increases
in urine, this is known as ketonuria.

• The overall picture of ketonemia and

ketonuria is called Ketosis.

• Both acetoacetate and β–hydroxy butyrate

are strong acids. They dissociate in blood and
release H+ ions, which lowers the pH
(between 7.0-7.2 but can be as low as 6.9).

• This decrease in blood pH as a result of

severe ketosis is called Ketoacidosis.
Acetone is volatile and can not be detected in the
blood. The odor of acetone may be detected in
the breath and also in the urine of a person who
has high level of ketone bodies in the blood e.g. in
severe diabetic ketoacidosis, while under normal
conditions, acetone formation is negligible.

– Diabetic Ketoacidosis is dangerous and may

result in coma and death if untreated.

– In severe Diabetes the KB levels in blood

may reach as high as 100 mg /dl and urinary
excretion may be as high as 500 mg/day.
 HYPERGLYCAEMIA AND DIABETES MELLITUS

• Hyperglycemia is the technical term for high blood glucose. It

may be due to reasons such as:

• Severe stress (usually transient)

• Intravenous infusion of glucose-containing fluids

• Diabetes mellitus or impaired glucose regulation

• Increased counter-regulatory hormones: Cushing’s syndrome,

Acromegaly, Thyrotoxicosis, Phaeocromocytoma

• Pancreas: Glucagonoma, Pancreatitis

 Diabetes Mellitus
• This syndrome is characterised by hyperglycaemia
due to deficiency of insulin and or insulin
resistance.
• It is defined on laboratory basis as FBS = or >
7.0mmol/l (on more than one occasion or once in the
presence of symptoms of Diabetes) and a
postprandial glucose (RBS) = or > 11.1 mmol/l
confirms the diagnosis of DM (WHO)
• Sometimes OGTT is used to establish diagnosis of
DM where the patient is given anhydrous glucose
for ingestion.
• Normal fasting blood glucose 3.4-6.2mmol/l,
random blood glucose 3.4-11.1mmol/l

• Physiological glucose does not fall below 4.0

mmol/l or rise above 10 mmol/l (Renal
threshold value)

TYPES OF DM
• Two classes of diabetes: IDDM and NIDDM
 TYPE 1
 It is also called insulin-dependent diabetes. It used to be called
juvenile-onset diabetes.

 Essentially an autoimmune disease condition. It develops when the

body’s immune system destroys pancreatic beta cells, the only cells in
the body that make the hormone insulin that regulates blood glucose.

 This type of diabetes may be caused by a genetic predisposition.

 This form of diabetes usually strikes children and young adults,

although disease onset can occur at any age.

 Type 1 diabetes may account for 5% to 10% of all diagnosed cases of

diabetes.

 Risk factors for type 1 diabetes may include autoimmune, genetic, and
environmental factors.
 Clinical and metabolic features of dm type 1
• Hyperglycaemia

• Glycosuria: normal RF but renal threshold is exceeded

• Polyuria: Osmotic diuresis (increased urination due to

presence of certain substances[glucose] in the kidneys

• Polydipsia (excessive thirst): cerebral cellular

dehydration due to hyperosmolality 20 to hyperglycaemia

• Lipolysis: FFA rise, FFA are converted to acetyl coA &

ketones/re-esterified to form endo. TG incorporated
into VLDL
 TYPE 2
• It is the commonest type of DM accounting for about 90% to 95% of all
diagnosed cases of DM.

• Hyperglycaemia may be due to either peripheral insulin resistance or

delayed insulin action. It usually begins as insulin resistance, a disorder
in which the cells do not use insulin properly. As the need for insulin
rises, the pancreas gradually loses its ability to produce insulin.

• Type 2 DM is associated with older age (late onset), obesity, family

history of diabetes, history of gestational diabetes, impaired glucose
metabolism, physical inactivity, and race/ethnicity.

• Subjects may not require insulin (dietary) therapy unless

hyperglycaemia get out of control.

• Patients are less likely to develop ketoacidosis

• NB: It is increasingly being diagnosed in children and adolescents.
 3. Diabetes associated with other conditions

• Absolute insulin deficiency due to pancreatic

disease

• Drugs such as thiazide diuretics

4. Gestational diabetes – it is Carb.

intolerance with onset or first recognition
during pregnancy. Due to anti-insulin
hormones secreted by the placenta
(Oestrogen, prL, cortisol, prog. etc)
 OTHER VARIANTS OF DIABETES
Maturity onset diabetes of the young (MODY)
• Mody is Type 2 diabetes that develops in typically type 1 age
group.

• Four genetic defects have been identified, one of which is a

mutation on the glucokinase gene.

• Glucokinase catalyzes conversion of glucose to glucose-6-

phosphate and eventually the metabolites that stimulate the
insulin secretion in the β-cell.

• Thus glucokinase serves as the glucose sensor for the β-cell.

• The defect means that increased levels of glucose are

required to elicit the usual secretion of insulin.
 SYNDROME X / METABOLIC SYNDROME
• A cluster of risk factors for heart disease associated with insulin
resistance described by Gerald Reaven in 1988.
The presence of three or more of the following features:

• Obesity BMI >29 (weight in kg divided by height in meters squared).

Waist: (Men -ideal <94: high risk- >102cm) (Women -ideal< 80: high
risk- >88cm)

• Hypertension (BP≥ 130/85 mmHg)

• Dyslipidaemia: (hypertriglyceridemia, small dense LDL and low

HDL)
 Fasting plasma triglycerides more than 1.7 mmol/L
 Low Fasting plasma high-density lipoprotein (HDL) [M: <0.9
mmol/L, F: <1.2 mmol/L]
• Cholesterol: (male < 1.0 mmol/L, female < 1.3
mmol/L), High serum LDL

• Insulin resistance: When normal amount of insulin

fails to maintain glucose within normal limits. This is
often associated with type II diabetes mellitus.

• High fasting blood glucose

• vi. High fibrinogen levels
• vii. Hyperuricaemia
• The main sign of metabolic syndrome is
central obesity (also known as visceral, male-
pattern or apple-shaped adiposity),
overweight with adipose tissue accumulation
particularly around the waist and trunk.

• Associated conditions include hyperuricemia,

fatty liver (especially in concurrent obesity)
progressing to non-alcoholic fatty liver
disease, polycystic ovarian syndrome (in
women), erectile dysfunction (in men) etc.
 Laboratory Test for DM
• Urine glucose (glucosuria)
• Fasting Blood Glucose (FBG)
• Random Blood Glucose (RBG)
• Oral Glucose Tolerance Test (OGTT)
• 2 hour Post Prandial (2hrPP)
• Glycated haemoglobin (HbA1c).
ACUTE METABOLIC COMPLICATIONS OF DIABETES MELLITUS

The main complications are:

• Diabetes ketoacidosis

• Coma

• Hypoglycaemia
 Diabetes ketoacidosis
• Blood glucose conc. 20-40 mmol/l
• Developed in diabetics as a result of insufficient/ interruption
of insulin therapy.

• May be precipitated by severe physical stress (trauma,

infection) resulting in the production of the counter regulatory
hormones like cortisol.
• It is aggravated by vomiting leading to the following conditions

1. fluid loss ultimately from both compartments

2. decreased circulatory blood volume
3. decreased GFR

• The ketone bodies (acetoacetate, 3-hydroxy butyrate and

acetone) may give rise to metabolic acidosis

• Breathing is rapid and deep referred to as Kussmaul`s

breathing
Symptoms of DKA
• Diabetic ketoacidosis signs and symptoms often develop
quickly, sometimes within 24 hours. For some, these signs
and symptoms may be the first indication of having diabetes.

Symptoms include:
• Excessive thirst
• Nausea and vomiting
• Frequent urination
• Abdominal pain
• Weakness or fatigue
• Shortness of breath
• Fruity-scented breath
• Confusion
• High blood sugar level (hyperglycemia)
• High ketone levels in your urine
Causes of DKA
• Diabetic ketoacidosis is usually triggered by:
• A problem with insulin therapy. Missed insulin treatments
or inadequate insulin therapy can result in too little insulin in
the body, triggering diabetic ketoacidosis.

• An illness: An infection or other illness can cause the body

to produce higher levels of certain hormones, such as
adrenaline or cortisol. e.g Pneumonia and urinary tract
infections.

• Other possible triggers of diabetic ketoacidosis include:

• Physical or emotional trauma

• Heart attack
• Alcohol or drug abuse, particularly cocaine
 COMA

• Marked hyperglycaemia but no detectable

ketonaemia or acidosis.

• Plasma glucose is usually about 50 mmol/l

• Associated glycosuria produces osmotic diuresis

with severe water and electrolyte depletion

• Coma results due to cerebral cellular dehydration

• Occurs commonly in patients with type 2 DM

 Hypoglycaemia

• It is the commonest cause of coma in diabetics

• Results when plasma glucose levels fall below 2.2 mmol/l.

• Symptoms vary depending on onset and glucose

concentration

Mild- symptom include: hunger, gastric hyperactivity,

irritability

Severe symptoms e.g overdose of insulin include; mental

confusion, thickness of speech, drunkenness, convulsion or
coma

• Untreated chronic hypoglycaemia in children may lead to

 Causes of Hypoglycaemia

1. Insulinoma

2. Glucagon deficiency

3. Ethanol-induced hypoglycaemia etc.

Three conditions must be satisfied (Whipple’s triad) when

diagnosing hypoglycaemia
 Symptoms of hypoglycaemia

 Low blood glucose (<2.2 mmol/L)

 Prompt reversal of symptoms on administration of glucose

 Late Complication of DM
• Neuropathy: Diabetic neuropathies are a family of nerve
disorders caused by diabetes. People with diabetes can,
over time, develop nerve damage throughout the body
• Some people with nerve damage have no symptoms.

• Others may have symptoms such as pain, tingling, or

numbness i.e loss of feeling in the hands, arms, feet, and
legs.
• Nerve problems can occur in every organ system,
including the digestive tract, heart, and sex organs.

• Macroangiopathy (or accelerated atherosclerosis):

abnormal wall of large blood vessels such as thickening of
the basement membrane due to poor glyceamic control, leads
to premature coronary heart disease
 Late Complication of DM

• Retinopathy: may cause blindness due to

vitreous haemorrhage from proliferating
retina vessels & maculopathy as a result of
exudates from vessels or oedema affecting
the macula

• Nephropathy (kidney disease): leads to

renal failure.
 MONITORING OF DM

• Blood glucose

• Urine glucose (glucosuria)

• Glycated haemoglobin (HbA1c). This is expressed as a

percentage of total blood haemoglobin conc. and gives a
retrospective assessment of the mean plasma glucose conc.
during the preceeding 6-8 weeks.

• Spuriously low values can be seen in chronic haemolysis

and in patients with conditions like SCD and G6PD
deficiency. Fructosamine is a better test in such conditions.

• Fructosamine is the glycosylation of albumin. It has a

shorter half-life i.e. 2 to 3 weeks. Ref (1.61-2.68 mmol/L)
 MANAGEMENT OF DM
• Education

Anti-Diabetic Drugs
• Insulin administration

• Oral medications e.g sulphonylureas, biguanides etc.

Sulphonylureas
• • These drugs were discovered by chance when it was noted that a
sulphonamide derivative used for treating typhoid, caused marked
hypoglycaemia
Examples include glibenclamide and chlorpropamide.

Biguanides
• • Improves insulin sensitivity by increasing glucose utilisation by skeletal
muscle and also decreases hepatic gluconeogenesis.

• Metformin is the most widely known and used drug in this class and is
frequently prescribed to overweight patients as it also promotes weight loss.