ENZYMES – WHY STUDY THEM

•Enzymes “make life on earth possible”

•Almost all biochemical reactions are enzyme catalysed.

•All enzymes (except ribozymes) are proteins.

•Many diseases(inborn) are due to genetic abnormalities in synthesis

of enzymes.

•When cell is injured(due to decreased blood supply or inflammation)

enzymes leak into the plasma which helps in diagnosis and
management of diseases.

•Enzymes are used as therapy

•Coenzymes / cofactors
 Discovery:

1. Late 1700s- digestion of meat by secretions of the stomach.

2. 1800s- conversion of starch to sugar by saliva and plant
extracts.
3. Louis Pasteur (1850s)- fermentation of sugar into alcohol by
yeast is catalyzed by “ferments”.
~ ferments are inseparable from living yeast cells- “vitalism”
4. Edward Buchner (1897)- yeast extracts could ferment sugar to
alcohol.
~ fermentation was promoted by molecules that
continue to function when removed from cells.
5. Frederick W. Kühne- called these molecules “enzymes”.
6. James Sumner (1926)- isolation and crystallization of
urease. ~ postulated that “all enzymes are proteins”
7. John Northrop and Moses Kunitz (1930s)-
crystallized pepsin, trypsin, and other digestive
enzymes.
~ all found to be proteins
8. J. B. S. Haldane (1850s)- wrote “Enzymes”
~ weak bonding interactions between an enzyme
and its substrate might be used to catalyze a
reaction.
9. Late 20th century- purification and elucidation of the
structure and mechanism of many enzymes.
 DEFINATION

Soluble, colloidal, organic catalysis(bio) produced by living

cells but also capable of acting independent of cells.

All enzymes with one exception are protein in nature

(RIBOZYME).

Essentially all biochemical reactions are enzyme catalysed.

 Extraordinary catalytic power of enzymes

1. High degree of specificity for their substrates

2. Accelerate chemical reactions tremendously
3. Function in aqueous solutions under very mild
conditions of temperature and pH
 terms
INTRACELLULAR ENZYMES
Glycolytic enzyme – cytosol
TCA enzyme - mitochondria
EXTRACELLULAR ENZYMES
Digestive enzymes

ZYMASE ZYMOGEN

SUBSTRATE PRODUCT

IRREVERSIBLE REVERSIBLE

MONOMERIC - ribonuclease
OLIGOMERIC – LDH
MULTIENZYME – Py Dh, FA synthetase
special pocket or cleft called the active sites.
 Chemical nature

All enzymes are proteins (ex: ribozymes)

But all proteins are not enzymes.
Enzymes show properties of proteins

•Unique structure of amino acid

•Ppt`d by usual protein ppting agent
•Non dialyzable
•Amphoteric
•Isoelectric point
•Show electrophoretic mobility
•Nitrogen content about ~16 %
•Sensitive to alteration in pH and temp.
 Activation energy or Energy of Activation:

• All chemical reactions require some amount of energy to

get them started.
OR
• It is First push to start reaction.

This energy is called activation energy.

It varies from one reaction to another

In an ordinary chemical reaction it is provided by heating a

reaction mixture

Enzyme accelerate reaction by lowering energy of

activation of reacting molecule
i.e. the transition state is achieved by the reacting molecule
easily in the presence of the enzyme.

Enz-Sub Complex Product

 Classification
• According to the International union Of
Biochemistry an enzyme name has two parts:

-First part is the name of the substrates for the

enzyme.
-Second part is the type of reaction catalyzed by
the enzyme. This part ends with the suffix “ase”.

Example: Lactate dehydrogenase

 EC (Enzyme commission)(IUB) number
1961 (with the latest update in 1992)

EC numbers are four digits,

• “a” is the class (broad function),
• “b” is the subclass (group involved),
• “c” is the sub-subclass (other functional requirement),
• “d” is the sub-sub-subclass (specific enzyme).

Example: for Alcohol:NAD+oxidoreductase

EC number is 1.1.1.1
Example:

Hexokinase: EC: 2.7.1.1

2: - Class i.e transferase

7:- Subclass i.e Transfer of phosphate
1:- SubSubClass i.e Alcohol as phosphate acceptor
1:- Specific enzyme name i.e: Hexokinase
 CLASSIFICATION OF ENZYMES

Enzyme commission of I.U.B:

Six major classes based on the type of reactions catalyzed

Class 1: Oxidoreductases
Class 2: Transferases
Class 3: Hydrolases
Class 4: Lyases
Class 5: Isomerases
Class 6: Ligases
E.C.1 Oxidoreductases

Catalysing oxidation reduction reaction where one

substrate is oxidized and other is reduced

Ascorbic acid oxidase,

Cytochrome oxidase, Tyrosinase
Glucose oxidase, L amino acid dehydrogenase,
Xanthene dehydrogenase
Lactate Dehydrogenase with NAD
Glucose 6 phosphate dehydrogenase with NADP
 E.C.2 Transferase
Catalysing transfer of substances without involving
oxidation reduction reaction

• Transaminases
• Transmethylase
• Transpeptidase
• Phosphotransferase
• Acetyl transferase
 E.C.3 Hydrolase
Catalysing hydrolytic breakdown of different bonds.
Most of the GIT enzymes belong to this class

• Amylase, maltase, lactase, sucrase

• Lipase, cholesterol esterase
• Phospholipase, carboxypeptidase
• Phosphatases
 E.C.4 Lyases
Catalysing reactions in which groups are removed
without hydrolysis leaving a double bond or add groups
to already existing double bonds.

• Pyruvate decarboxylase
• Fumarase
 E.C.5 Isomerase
Involved in inter conversion of pair of isomeric
compounds

• Phosphoglucomutase
• Phosphohexose isomerase
• Retinene isomerase
 E.C.6 Ligases
Catalyze reactions in which linking together of two
molecules occur coupled with the breakdown of a high
energy phosphate bonds like ATP, GTP

• Acetyl CoA syntethase

• Succinyl CoA synthetase
• Pyruvate carboxylase
• Acyl CoA Synthetase
 EC-7: Translocases (A new EC Class: 2018)

Translocases catalyze the movement of ions or

molecules across membranes or their separation within
membranes.
. Eg:
Ornithine Translocase (Na KATPase)
 Enzyme Specificity

• Enzymes have varying degrees of specificity

for substrates
• Enzymes may recognize and catalyze:
- a single substrate
- a group of similar substrates
- a particular type of bond
 Types of enzyme specificity
Type Reaction Example
Absolute Catalyse one type of Urease- hydrolysis
reaction for a single of urea
substrate
Group Catalyse one type of Hexokinase adds
reaction for a similar phosphate to
substrate hexoses

Linkage Catalyse one type of Chymotrypsin,

(bond) reaction for a specific hydrolysis of
type of bond peptide bonds
Stereo Specific for L/D forms L amino acid, D
Sugars
Mechanism of enzyme action
 Enzyme Catalyzed Reactions (Michelis Menton)
• When a substrate (S) fits properly in an active site, an
enzyme-substrate (ES) complex is formed:
E + S  ES
• Within the active site of the ES complex, the reaction
occurs to convert substrate to product (P):
ES  E + P
• The products are then released, allowing another substrate
molecule to bind the enzyme
- this cycle can be repeated millions (or even more) times
per minute
• The overall reaction for the conversion of substrate to
product can be written as follows:
E + S  ES  E + P
 Lock-and-Key Model(Fischer)
- the active site has a rigid shape
- only substrates with the matching shape can fit
- the substrate is a key that fits the lock of the active site

• This is an older model and does not work for all enzymes
 Induced Fit Model (Koshland)
- the active site is flexible, not rigid
- the shapes of the enzyme, active site, and substrate
adjust to maximize the fit, which improves catalysis
- there is a greater range of substrate specificity
• This model is more consistent with a wider range of
enzymes
 Factors affecting enzyme activity (velocity of enzy)
Enzyme activity: expressed as IU
A unit of activity is defined as the amount of enzyme that will produce
1 micromoles of product per minute at 250 C under standardized
conditions
1. Conc of enzyme
2. Conc of substrate
3. Temperature
4. H+ ion conc or pH
5. Time
6. Product of reaction
7. Coenzyme conc
8. Enzyme activator (Cofactor)
9. Effect of oxidation
10.Effect of radiation
11.Enzyme inhibitors
CONC OF ENZYME
Velocity is directly proportional to enzyme concentration
(subject to availability of substrate)
Initial reaction rate / arbitrary

For endpoint estimation

units

Enzy conc
CONC OF SUBSTRATE

Zero order kinetics

1st phase
1st order kinetics
Half enzy saturated with subs
Subs. conc required for
attaining half the maximum
velocity (activity) of the
enzyme
Km

•Substrate conc at half maximal velocity

•50% of enzyme molecule saturated with substrate
•Independent of enzyme concentration
•Characteristic feature (signature ) of an enzyme
•Denotes affinity of enzyme to the substrate
•More the affinity less the dissociation
THE MICHAELIS-MENTON EQUATION

Velocity (V1) is expressed according to MM Equation

Describes the behaviour of

many enzymes as substrate
conc is varied and also helps
to determine km for any
enzyme.

LIMITATIONS OF MM MODEL
SIGMOIDALCURVE FOR Hb
Indicates cooperative binding
of substrate to multiple sites
Lineweaver- burk plot
 EFFECT OF TEMPERATURE

Q10= Temperature coefficient

Reaction velocity doubles with every 10O C
ENZYME ACTIVITY DEPENDS ON PH
 COENZYME
Specific non protein low molecular weight, dialysable, heat stable,
organic compound

Same compound can act as coenzyme for more than one enzyme.

Apoenzyme portion determine enzyme specificity.

Most water soluble vitamins are converted in the body into coenzyme.

Coenzyme requiring enzymes are oxidoreductases, grp transfer,

isomerisation and reaction forming covalent bond.

Enzymes not requiring coenzymes – lytic and hydrolytic reactions

Coenzymes can be regarded as second substrate or a cosubstrate –

chemical changes in a coenzyme exactly counter balance those taking
place in substrate
 Coenzyme classification based on functional
characteristics

1. Coenzyme involved in group transfer or atoms other than

hydrogen
a. ATP and related compound e. Sugar phosphates
b. CoA f. TPP
c. B6 phosphate g. Biotin
d. Folate enzyme h. Vitamin B12

2. S-adenosyl methionine

3. Coenzyme involved in transfer of hydrogen atom

NAD-NADP, FAD-FMN, Lipoic acid, CoQ, Cytochromes
Vitamins of B complex group acting as co-enzymes
vitamins active form (co-enzyme)

Thiamine Vitamin B 1 TPP (thiamine pyrophosphate)

Riboflavin Vitamin B 2 FMN, FAD
Niacin Vitamin B 3 NAD,NADH
Pantothenic acid Vitamin component of coenzyme A
B5
Pyridoxine Vitamin B 6 PLP (pyridoxal phosphate)
Biotin Biotin
Folic acid THF (Tetrahydrofolate)
Cobalamine Vitamin B 12 cobamide
 Coenzyme involved in transfer of groups other than
hydrogen

•ATP and related compounds (UTP,CTP,GTP,cAMP,CDP,GDP)

•Coenzyme A- involved in transfer of acyl group

Lipmann in 1947

Pantothenic acid-  alanine- mercaptoethylamine-two phosphates-3`phophoadenosine unit

Essential for FA and cholesterol synthesis as acetyl CoA

Eg: Thiokinase, Thiolase, Acyl Transferase
•Pyridoxal phosphate
Active form of B6 or pyridoxine
Eg: Transamination (AST, ALT), Decarboxylation

•Lipoic acid
Eight carbon disulphide containing carboxylic acid i.e
attached to dihydrolipoyl transacetyalse by an amide bond.
Pyruvate DH and α KG DH

•Thiamine pyrophosphate (TPP)

Eg: Pyruvate DH,  keto glutarate, transketolase
•Tetrahydro folate (FH4)(Folic acid)
Required for the transfer of single carbon units such as
formyl, methyl, methylene groups
In biosynthesis of amino acids and purine

•Biotin
Required for carboxylation reactions
Eg: Pyruvate carboxylase
Propionyl carboxylase
Acetyl CoA Carboxylase
•Cobamide or B12
Cobalt containing B complex vitamin
Methyl malonyl CoA to Succinyl CoA

•S-adenosyl methionine
-Transmethylation reaction
Metalloenzymes:
Metal tightly bound to enzyme

Metal activated enzyme:

Loosely bound, can be removed by dialysis
Activators (Cofactors, metalloenzymes) Positive Modifiers
Element Enzyme
Zinc Carbonic anhydrase, Alcohol dehydrogenase
Molybdenum Xanthine oxidazse, Isocitrate Dehydrogenase
Iron Catalase, Peroxidase
Copper Ascorbic acid oxidase, tyrosinase, Diamine oxidase
Magnesium Phosphorylase, Glucokinase, Thiokinase
Potassium Arginine synthetase, Pyruvate kinase
Calcium Clotting enzyme
Chloride Salivary enzyme
Cu, Zn, Mn Superoxide dismutase
Fe, Cu Cytochrome oxidase
Mg Na K Mg Na K ATPase
 Enzyme inhibition
Enzyme inhibitors are molecular agents that interfere with
catalysis, slowing or halting enzymatic reactions.

Enzyme activity can be regulated by the following methods:

1. Competitive and noncompetitive inhibitions
2. Allosteric control
3. Multiple forms of enzymes (isozymes)
4. Reversible covalent modification
5. Proteolytic activation, and
6. Controlling the amount of enzyme present.
-- reversible
-- inhibitor (usually) structurally similar to substrate
-- inhibitor competes directly for substrate binding to
active site (mutually exclusive binding)
-- effects can be overcome by increasing substrate
concentration
-- Km is increased, Vmax not changed
Reaction Enzyme Inhibitor
Succinic acid to Succinate DH Malonic acid
fumaric acid
Citrate to isocitrate Aconitase Fluroacetate
 Clinically used competitive inhibitors
Drug Enzyme Clinical use
Allopurinol Xanthine oxidase Gout
Penicillin Transpeptidase Antibiotic
Sulphonamide Pteroid syntethase Antibiotic
Trimethoprim FH2 reductase Antibiotic
Methotrexate FH2 reductase Cancer
5fluorouracil Thymidylate synthase Cancer
6 mercaptopurine Adenylosuccinate synthase Cancer
Acyclovir DNA polymerase Virus infection
Lovastatin HMG Co A reductase Cholesterol
lowering
Neostigmine Ach- esterase Myasthenia
Alpha methyl dopa Dopa decarboxylase Hypertension
 / Non-competitive

--Binds to site on enzyme remote from active site.

--No structural resemblance to substrate
-- Causes conformational change in enzyme that prevents
conversion of substrate to product, but does not
prevent substrate binding to enzyme.
Km is not changed, Vmax is reduced
Non-competitive

Cyanide inhibits cytochrome oxidase

Fluoride inhibits enolase
BAL - SH groups
Diisopropyl flurophosphate inhibits actelycholine esterase
Metal poisons
 Competitive Non competitive
inhibition inhibition
Acting on Active site May or may not
Structure of Substrate analog Unrelated
inhibitor molecule
Inhibition is reversible Generally
irreversible
Excess substrate Inhibition relieved No effect
km Increased No change
Vmax No change Decreased
Significance Drug action Toxilogical
-- Cannot bind to free enzyme.
-- Binds only to enzyme-substrate complex (ES).
* substrate binds directly to inhibitor, or
* substrate induces conformational change required for
inhibitor binding.
-- Once bound, inhibitor prevents enzyme from converting
substrate to product.
Vmax and Km decreased

Eg: Inhibition of placental

alkaline phosphatase
(Regan isoenzyme) by
phenylalanine
Suicide inhibition - mechanism-based inactivation, Irreversible inhibition

The inhibitor makes use of the enzyme’s own reaction mechanism to

inactivate it.

The structural analog is converted to a more effective inhibitor with

the help of the enzyme to be inhibited.

Allopurinol which is oxidized by xanthine oxidase to alloxanthine

that is a strong inhibitor of xanthine oxidase.

Aspirin - Arachidonic acid is converted to prostaglandin by the

enzyme Cyclo-oxygenase. Aspirin acetylates a serine residue in the
active center of cyclo-oxygenase, thus prostaglandin synthesis is
inhibited, and so inflammation subsides.

Ornithine decarboxylase
Feedback inhibition / End product inhibition

Delta-Amino levulinic acid synthase by heme

AMP in purine biosynthesis

Aspartate transcarbamolase by CTP

Delta-Amino levulinic acid synthase (HEME)

Induction – barbiturates

Inhibitor – heme (decreases reaction, enzyme activity)

Repressor – blocks the synthesis of enzyme

Feedback Inhibition
Activity of the enzyme is inhibited by the product of the reaction.

Inhibition of hexokinase by glucose-6-phosphate is an example.

Glucose + ATP → Glucose-6-phosphate + ADP

Succinyl CoA + Glycine --------- delta aminolevulinic acid (ALA)

(The end product, heme will allosterically inhibit the ALA synthase).
 Allosteric regulation
Allosteric enzyme has one catalytic site where the
substrate binds and other separate site where the
modifier binds (allo- other).

Substrate and allosteric binding site may or may not be

adjacent.

The inhibitor is not a substrate analog

Partially reversible, when excess substrate is added

 Allosteric regulation

Most allosteric enzymes possess quaternary structure

Molecule (Positive / negative modifier) can increase or

decrease the activity of allosteric enzyme.

Positive cooperativity – Binding of substrate to one of

the subunits of the enzyme may enhance substrate
binding by other subunits.

Negative cooperativity-
Salient features of allosteric inhibition

1. The inhibitor is not a substrate analog.

2. It is partially reversible, when excess substrate is
added.
3. Km is usually increased.
4. Vmax is reduced.
5. The effect of allosteric modifier is maximum at or near
substrate concentration equivalent to Km.
6. Most allosteric enzymes possess quaternary structure.

They are made up of subunits, e.g. Aspartate

transcarbamoylase has 6 subunits and pyruvate kinase
has 4 subunits.
Enzymes Allosteric Allosteric
inhibitor activator
Phosphofructokinase ATP, Citrate AMP, F26P
ALA synthase Heme
Aspartate CTP ATP
transcarbamoylase
HMG CoA reductase Cholesterol
Pyruvate carboxylase ADP Acetyl CoA
AcetylCoA carboxylase Acyl CoA Citrate
Citrate synthase ATP
Carbamoyl phosphate NAG
synthetase I
Carbamoyl phosphate UTP
synthetase II
Regulatory enzymes are modulated in a variety of ways

Allosteric enzymes function through reversible

noncovalent binding of regulatory compounds
called allosteric modulators or allosteric effectors.

Other enzymes are regulated by reversible covalent

modification.
The activity of enzymes may be increased or decreased by
covalent modification.

Either addition of a group to the enzyme protein by a

covalent bond; or removal of a group by cleaving a
covalent bond.

Zymogen activation by partial proteolysis is an example

of covalent activation.

cAMP –Glycogen metabolism

 Examples of covalent modification

Enzyme Phosphorylated enzyme

Acetyl CoA carboxylase Inactive
Glycogen synthase Inactive
Pyruvate DH Inactive
Glycogen phosphorylase Active

And many more

Induction / repression

Lac operon
ENZYMES USED AS THERAPEUTIC AGENTS
Enzyme Therapeutic application
Asparaginase Acute lymphoblastic leukemia
Streptokinase To lyse intravascular clot
Urokinase To lyse intravascular clot
Recombinant tissue prothrombin Lysis of clot, especially for
activator (rtPA) cerebrovascular thrombolysis
Streptodornase DNAse applied locally
Pancreatin (trypsin and lipase) Pancreatic insufficiency; oral
administration
Papain Anti-inflammatory
Alpha-1-antitrypsin AAT deficiency; emphysema
ENZYMES USED IN LABORATORY MEASUREMENTS

Enzyme Used for testing

Urease Urea
Uricase Uric acid
Glucose oxidase Glucose
Peroxidase Glucose; Cholesterol
Hexokinase Glucose
Cholesterol oxidase Cholesterol
Lipase Triglycerides
Horse radish peroxidase (HRP) ELISA
Alkaline phosphatase ELISA
Restriction endonuclease Southern blot; RFLP
Reverse transcriptase Taq polymerase Polymerase chain reaction (PCR)
 CLINICAL ENZYMOLOGY
ENZYMES OF DIAGNOSTIC SIGNIFICANCE

Rona first : serum lipase estimation

Functional Plasma Enzymes:

Secreted in the plasma and function in the plasma
Eg: Blood coagulation and dissolution enzymes
Lipoprotein lipase

Non-Functional Plasma Enzymes:

No function in the blood
Substrate present elsewhere.
Low normal levels in plasma
Increase indicates tissue destruction, leakage from living or
dead or dying cells.
Enzyme Unit : One international Unit (IU)

Amount of enzyme that will convert one

micromole substrate per minute per liter of
sample into product
 Enzymes of diagnostic significance

ENZYME NORMAL VALUE SIGNIFICANCE

SGOT(AST) 8-40 U/L MI – 2nd to rise in 3-4
days
Hepatic disease
SGPT (ALT) 6-35 U/L Acute hepatitis
Alkaline 4-11 KAU Obstrutive jaudice
phosphatase 40-125 U/L Bone disease – rickets,
osteoblastoma, Ca bone
Inf hepatitis.
Carcinoma
Hodgkins lymphoma
Acid phosphatase 2.5-12 U/L Prostate cancer
PSA
Amylase 80- 180 s Acute pancreatitis
units Mumps
Obstruction of duct
Acute abd conditions
Lipase 0.2 -1.5 U/L Acute pancreatitis
LDH 100-200 U/L Myocardial Infarction
(5-10 times, 4-7 days)
Hemolytic anemia
Hepatocellular damage
Muscular dystrophy,
Carcinoma, Leukemia
Creatine kinase 15-100 U/L CK-MB within 3 hours of
MI
Muscle disorders- CKMM
Brain disorders - CKBB
Glucose 6 In RBC 125- Haemolytic anemia
phosphate 250 U/
dehydrogenase
Glucose-6-phosphate Dehydrogenase (GPD)
HMPS

used for production of NADPH- preserving the RBC integrity.

Drug-induced hemolytic anemia:

In the GPD deficient individuals, RBC lifespan may be reduced, but
there will be no disease manifestations.
But when certain drugs (aspirin, mepacrine, primaquine, sulpha)
are taken by such individuals, there will be sudden damage to
RBCs.
This drug-induced hemolytic anemia is characteristic of GPD
deficiency.

Fava beans may also induce hemolytic anemia which is called

favism.
Carrier State has Biological Advantage
The gene for GPD is located in X-chromosome.

In heterozygous condition, where one gene is abnormal and the

allelic form is normal, the GPD level in RBC is half the normal
value.

GPD deficiency seems to protect the person from falciparum

malaria. The malarial parasites require NADPH for optimal growth.
Ceruloplasmin 3-58 mg% Cirrhosis, Bacterial
infection, pregnancy
5` Nucleotidase 2-17 IU/L Acute liver disease
Obstructive jaundice
Tumours
Gamma glutamyl 10-30 U/L Infective hepatitis
transferase Prostate cancer
Alcoholism
Obstructive jaundice
Enzyme Patterns (Enzyme Profiles) in Diseases
I. Hepatic diseases
1. Alanine aminotransferase (ALT): Marked increase in
parenchymal liver diseases
2. Aspartate aminotransferase (AST): Elevated in parenchymal liver
disease
3. Alkaline phosphatase (ALP): Marked increase in obstructive liver
disease
4. Gamma glutamyl transferase (GGT): Increase in obstructive and
alcoholic liver
Enzyme Patterns (Enzyme Profiles) in Diseases
II. Myocardial infarction
1. Cardiac troponins (CTnT and CTnI). (These are not enzymes, but
are specific and sensitive and elevated very early in MI).
2. Creatine kinase (CK-MB): CK-MB isoenzyme is specific

III. Muscle diseases

1. Creatine kinase (CK-MM): Marked increase in muscle diseases.
2. Aspartate aminotransferase (AST): Increase in muscle disease;
not specific
3. Aldolase (ALD): Earliest enzyme to rise, but not
specific
IV. Bone diseases
1. Alkaline phosphatase (ALP) Marked elevation in rickets and
Paget’s disease

V. Prostate cancer
1. Prostate specific antigen (PSA): Marker for prostate cancer. Mild
increase in benign prostate enlargement
2. Acid phosphatase (ACP): Marker for prostate cancer. Metastatic
bone disease especially from a primary form prostate. Inhibited by
L tartrate.

VI. Pancreatic disease

1. Amylase: Marker for acute pancreatitis and inflammation of
salivary glands
2. Lipase: Marker of pancreatitis, more specific than amylase
ISOENZYMES OR ISOZYMES

Tissue 1 Tissue II Tissue III

A B A B A B

Physically distinct forms of the same enzyme found in

different tissues BUT catalyses the same chemical reaction
and show different properties like:
Electrophoretic mobility
Rate of reaction
Immunological behaviour
Optimum pH
Structure
LDH (Lactate dehydrogenase)

PA LA
NAD NADH

*Tetramer with four subunits of Two

*H(Heart) or M(Muscle) polypeptide chains
•Five combinations
•With different electrophoretic mobility
•LDH-1 moves fast, LDH-5 Slowest
Formation of isoenzymes of lactate dehydrogenase.
Electrophoresis pattern of LDH isoenzymes.
Clinical significance:

Total LDH increases in damage to myocardium / liver / muscle

LDH1 & 2 increase in myocardial infarction(3-6 days later)

LDH 4&5 in acute liver hepatitis

LDH 5 in breast cancer, CNS prostate malignancies

LDH 2,3 in leukemias

 Isoenzymes of Creatine kinase:

•Dimer: Two subunits B(brain) and M(muscle)

•Three combinations BB,MB,MM called as CK1,2,3.
•CK-MB important since it rises the earliest, within 3-4
hours of infarction. (reaches peak in 24 hours)
•Two more atypical
Isoenzymes isoenzymes
of alkaline of CK reported: CK-
phosphatase:
macro isoenzymes
•Many and CK Mi(mitochondria)
•Major in serum are from liver, brain, bone and intestine.
•-1 ALP increases in obstructive jaundice.
•-2 ALP heat labile at 65oC increases in hepatitis.
•Pre  ALP in bone disease.
Isoenzymes of aldolase A and B