Isoenzymes

1 Introduction.- Nomenclature of multiple forms of enzymes. Numbering of isoenzymes.- 2 Techniques for the Separation of Isoenzymes.- Preparation of materials for isoenzyme separation. Electrophoretic techniques: general considerations; paper electrophoresis; starch-block electrophoresis; cellulose acetate electrophoresis; agar-gel electrophoresis; starch-gel electrophoresis; polyacrylamide-gel electrophoresis; column electrophoresis; iso-electric focusing. Chromatographic techniques:ion-exchange celluloses. Gel filtration.- 3 Detection and Determination of Isoenzymes.- Elution techniques: NAD- and NADP-dependent dehydrogenases; esterases; peroxidases. Detection of isoenzymes in electrophoretic media: ‘enzymoelectrophoresis’; tetrazoliumstaining techniques; ‘nothing dehydrogenase’ effect; staining techniques for esterases.- 4 The Chemical Nature of Isoenzymes.- Sub-unit structure: lactate dehydrogenase; creatine kinase;aldolase; glucose 6-phosphate dehydrogenase; tryptophan synthetase; catalase; caeruloplasmin. Presence of charged groups in isoenzyme molecules: alkaline phosphatase. Polymerization as the source of enzyme heterogeneity: cholinesterases; amylase. Conformational isomerism: malate dehydrogenase. Differences in amino-acid sequence.- 5 Enzyme Multiplicity in the Glycolytic Pathway and the Pentose—Phosphate Cycle.- Hexokinase: detection of hexokinase after zone electrophoresis; hexokinase isoenzymes in the erythrocyte; hexokinase in other tissues. Phosphoglucomutase. Glucose 6-phosphate dehydrogenase: demonstration of glucose 6-phosphate dehydrogenase heterogeneity; chemical structure and properties of glucose 6-phosphate dehydrogenase; genetic variants of human erythrocytic glucose 6-phosphate dehydrogenase; glucose 6-phosphate dehydrogenaseheterogeneity in other tissues. 6-Phosphogluconate dehydrogenase: heterogeneity in human erythrocytes; 6-phosphogluconate dehydrogenase isoenzymes in other species; structure of 6-phosphogluconate dehydrogenase isoenzymes. Phosphoglucose isomerase. Aldolase: classification of the aldolases;electrophoretic studies; fructose 1-phosphate aldolase. Glyceraldehyde 3-phosphate dehydrogenase. ?-Glycerophosphate dehydrogenase. Phosphopyruvate hydratase (enolase). Pyruvate kinase.- 6 Lactate Dehydrogenase Isoenzymes.- Distribution. Chemical and kinetic properties: substrate specificities and affinities; effect of pH variation; effects of inhibitors; reactions with coenzyme analogues; effect of temperature. Lactate dehydrogenase isoenzymes in developing tissues. Genetic variants of lactate dehydrogenase isoenzymes. Diagnostic applications of lactate dehydrogenase isoenzymes:myocardial infarction; liver diseases; malignant diseases;diseases of muscle; diseases of joints: anaemia; pregnancy;renal diseases; non-electrophoretic diagnostic techniques:serum 2-hydroxybutyrate dehydrogenase in diagnosis; relative heat-stability test; solvent-precipitation techniques; selective absorption techniques. Summary.- 7 Multiple Molecular Forms of Other Oxidoreductases.- Malate dehydrogenase: mitochondrial and supernatant malate dehydrogenase isoenzymes; biological significance of malate dehydrogenase isoenzymes; malate dehydrogenase isoenzymes in mammalian tissues; malate dehydrogenase isoenzymes in insect development; malate dehydrogenase isoenzymes of the chicken embryo. Isocitrate dehydrogenase: isocitrate dehydrogenase isoenzymes in heart and liver diseases. Glutamate dehydrogenase. Alcohol dehydrogenase: alcohol dehydrogenases of insects; avian alcohol dehydrogenases; alcoholdehydrogenases in mammalian liver; alcohol dehydrogenase polymorphism in plants. Xanthine dehydrogenase. Miscellaneous dehydrogenases.- 8 Aminotransferase (Transaminase) Isoenzymes.- Separation of aspartate aminotransferase isoenzymes. Properties of aspartate aminotransferase isoenzymes. Clinical applications of aspartate aminotransferase isoenzymes. Aspartate aminotransferase isoenzymes in developing tissues. Alanine aminotransferase.- 9 Phosphatase Isoenzymes.- Alkaline phosphatase: distribution, paper electrophoresis; starch-block electrophoresis; agar-gel electrophoresis; starchgel electrophoresis; acrylamide-gel electrophoresis; electrophoresis on Sephadex G-200; cellulose acetate electrophoresis; ion-exchange chromatography; properties of alkaline phosphatases; substrate affinities; effects of inhibitors; heat inactivation; effect of neuraminidase; summary of catalytic properties of tissue alkaline phosphatases; immunochemical reactions; genetic studies; alkaline phosphatase isoenzymes in bacteria. Acid phosphatase; electrophoretic separation; chromatographic investigations; genetic variants of human acid phosphatases.- 10 Multiple Forms of Esterases.- Aliesterases: genetic variations of non-specific esterases; ontogenetic observations. Cholinesterases: plasma cholinesterases; genetic polymorphism of serum cholinesterases; Cholinesterase isoenzymes in other species. Arylesterases:serum arylesterases; tissue esterases. Esterases and isoenzymes.- 11 Miscellaneous Enzymes.- Arylamidase: arylamidase isoenzymes in man; arylamidases in other species. Alanine aminopeptidase. Creatine kinase: creatine kinase isoenzymes. Arginine kinase. Caeruloplasmin (copper oxidase). Catalase. Carbonic anhydrase. Amylase: serum and tissue amylases; urinary amylase; macroamylasaemia. Other enzymes occurring in multiple forms: glycogen Phosphorylase; adenylate kinase; adenosine deaminase; tyrosinase; ribonuclease; deoxyribonucleic acid (DNA) polymerase; peroxidase; hyaluronate lyase (hyaluronidase); ?-glycosidases.- 12 The Biological Significance of Isoenzymes.- Author Index.