ISOENZYME – LACTATE

DEHYDROGENASEby,ISHAQUE P KBIOCHEMISTRYPondicherry university

About…

• Isozymes also known as isoenzymes (or more generally as Multiple forms of enzymes) are the physically distinct forms of enzyme but catalyses the same chemical reaction.

• Enzymes that differ in structurally (amino acid sequence), electrophoretically, immunologically and site of occurrence.

• Different isoenzymes are often organ-specific and their determination may improve the specificity of enzyme tests.

• The heterogeneity of some isoenzymes is due to different protein subunits which are coded for by separate genes.  

Reasons For Isoenzyme……

• Synthesized from different genes.

(Eg: malate dehydrogenase in cytosol versus in mitochondria)

• Oligomeric forms of more than one type of subunits.

(Eg: lactate dehydrogenase)

• Different carbohydrate content.

(Eg: alkaline phosphatase)

Examples……

Creatine Phosphokinase(CPK):• CPK-1 (also called CPK-BB) is found mostly in the brain

and lungs.• CPK-2 (also called CPK-MB) is found mostly in the heart.• CPK-3 (also called CPK-MM) is found mostly in skeletal

muscle.

Hexokinase:• Hexokinase I/A.• Hexokinase II/B.• Hexokinase III/C .• Hexokinase IV, also referred to as glucokinase.

Alkaline phosphatase (AP, ALP, SAP) :• Liver-ALP (L-ALP).• Corticosteroid-ALP (C-ALP, only in dogs). • Bone-ALP (B-ALP).• Intestinal-ALP (I-ALP). 

Aspartate transaminase (AST/ASAT/AAT) or serum glutamic oxaloacetic transaminase(SGOT):

• GOT1/cAST, the cytosolic isoenzyme derives mainly from red blood cells and heart.

• GOT2/mAST, the mitochondrial isoenzyme is present predominantly in liver.

• "Lactate dehydrogenase (LDH) is an enzyme present in a wide variety of organisms, including plants and animals".

• Its Enzyme Comission number is EC 1.1.1.27 where;

– EC 1 = oxidoreductase.

– EC 1.1 = acting on the CH-OH group of the donor.

– EC 1.1.1 = With NAD or NADP as acceptor.

– EC 1.1.1.27 = L-lactate dehydrogenase.

• Thus, it is an oxidoreductase enzyme that reversible reaction of lactate(LA) to pyruvate(PA) accompanied by the interconversion of NADH and NAD+.

Lactate Dehydrogenase(LDH)

• In serum five physically distinct isoenzymes exist and are known as LDH-1, LDH-2, LDH-3, LDH-4, LDH-5.

• All these enzymes catalyze the same reaction of oxidation of LA to PA.

• A sixth, typical isoenzyme LDH has been found in male genital tissues, called LDHx.

• Each Isozymes protein is made up of four polypeptide subunits, thus each is a “tetramer”.

• Each tetramer consists of four sub-units or monomers each with mass of 36 kDa giving the tetramer the mass of 144 kDa.

• Each monomer consists of a peptide chain of 334 amino acids with its own active center.

• Each subunit maybe one of two types termed H and M.• The different isoenzymes contain H and M in different

proportions.

• The different form can be separated by electrophoresis.• The difference in electrophoretic mobilities due to

different electric charges on the isoenzymes due to difference in content of acidic and basic amino acids.

• The H gene is more strongly negatively charged than M gene due to higher number of acidic residues. Example:– LDH-1 has the highest negative charge and hence,

moves fastest during electrophoresis. It contains a higher proportions of Asp and glutamate than the other forms.

– LDH-5 is the slowest moving fraction.

Identification…….

• Rate of chemical reactions is catalyzed, the different isoenzymes may catalyzes the same reaction at different rates.

Example:

– Rate of oxidation of Hydroxy Butyrate is greater by LDH-1 and LDH-2, when compared with rate of oxidation of LDH-4 and LDH-5.

• The isoenzymes may have different physical properties also.

Example:

– LDH-4 and LDH-5 are easily destroyed by heat, whereas LDH-1 and LDH-2 are not, if heated up to about 60oc (“Heat-resistant”).

• Myocardial LDH (LDH-1) is found to be more heat stable than that of Hepatic LDH.

• Hepatic LDH (LDH-5) is inhibited by urea.

• The isoenzymes have different pH optima and Km values.• The isozymes have different Km values for pyruvate and

therefore are better suited for different environments inside the body.

• A pure tetramer of M subunit i.e. M4, has lower Km for pyruvate and is concentrated in skeleton muscles which are anaerobic.

• Therefore M4 promotes glycolysis by catalyzing the production of lactate from pyruvate quickly and efficiently.

• On the other hand pure tetramer of H subunit i.e. H4, has greater Km for pyruvate and is more concentrated in heart muscles which is aerobic.

Clinical Significance……

• Normal serum LDH level is 60 – 250 IU/L.

• After damage to either of tissues viz; myocardium or liver, total serum LDH is increased and it may be useful to know the origin of the enzyme increase.

• In normal serum, LDH-2 is predominent isoenzyme and the slowest peak of LDH-5 is rarely seen.

• After myocardial infarction, the faster isoenzymes LDH-1 and LDH-2 predominate.

• In acute viral hepatitis, the slowest isoenzymes LDH-4 and LDH-5 predominate.

• Total serum LDH is frequently elevated in neoplastic diseases.

• In malignancies, isoenzyme pattern shifts towards slower migrating zone, there is increase usually LDH-3, LDH-4 and LDH-5.

• An increase in LDH-5 is seen in Breast carcinoma, malignancies of CNS, prostatic carcinoma.

• In leukemia, rise more in LDH-2 and LDH-3.

• Malignant tumors of testes and ovary shows rise in LDH-2, LDH-3 and LDH-4.

Reference………

• http://www.ukessays.com/essays/chemistry/the-lactate-dehydrogenase.php.

• http://www.nlm.nih.gov/medlineplus/ency/article/003499.htm.

• http://www.umm.edu/ency/article/003499.htm.

• http://www.sciencedirect.com/science/article/pii/030519789390014I.

• http://en.wikipedia.org/wiki/Lactate_dehydrogenase.