4 You are supposed to know … Enzymes – main features, propertiesCoenzymes – structures, functionsEnzyme kineticsEnzyme activity
5 Isoenzymes – General Features Genetically determined differences in primary structureCatalyze the same reactionMay have different subcellular distribution (cytoplasm × mitochondria)May have different tissue distributionMay be combined from more subunits (quarternary structure)May differ in kinetic properties (KM)Usually are determined by electrophoresis
6 Explain the terms: proenzyme, isoenzyme, isoform Q. 2 (p. 27)Explain the terms: proenzyme, isoenzyme, isoform
7 A. 2Proenzyme (zymogen) – inactive form of enzyme that becomes active after partial proteolysis example: pepsinogen pepsinIsoenzyme – see previous pageIsoform – more general term, includes true isoenzymes and pseudoisoenzymes (posttranslational variations)
8 Lactate dehydrogenase (LD) TetramerTwo different chains (H - heart, M - muscle)Five isoenzymes: LD1 (H4), LD2 (H3M), LD3 (H2M2), LD4 (HM3), LD5 (M4)Widely distributed in bodyTotal activity determination – nonspecific findingLD1 + LD2 ….. marker of myocardial infarction (MI)Today is LD assay considered out-of-date
9 Creatine kinase (CK) Dimer Two different chains (M – muscle, B – brain)Three isoenzymes: MM (muscle), MB (heart), BB (brain)Major isoenzyme in blood is MM (95 %)MB form in blood: 0 – 6 %BB in blood: traces (BB cannot pass across blood-brain barrier)MB isoenzyme …. excellent marker of myocardial infarction
10 Intracellular enzymes Enzymes in BloodFeaturePlasmatic enzymesSecretory enzymesIntracellular enzymesExampleSource organFunction incoag. factorsliverbloodamylase, lipasepancreasGITAST, …variouscellsEnzyme activity in blood after source organ damage?
11 Intracellular enzymes Enzymes in BloodFeaturePlasmatic enzymesSecretory enzymesIntracellular enzymesExampleSource organFunction incoag. factorsliverbloodamylase, lipasepancreasGITAST, …variouscellsEnzyme activity in blood after source organ damage
12 Q. 6Why are low activities of cellular enzymes detected even in serum of healthy people?
13 A. 6 Low activities of intracelular enzymes in extracelular fluid (blood plasma, serum)are the consequenceof physiological cell disintegration.
14 Main Tissue Distribution of Enzymes ASTALTLDCKGMTALPACPAMSLPSCHSliver, myocardlivernot specificmyocard, musclesbiliary tract, bonesprostatepancreas
15 Intracellular Location of Enzymes CytoplasmMitochondriaGolgi complex, ERLysosomeMembraneLD, ALT, 30 % AST70 % ASTCHS, AMSACPGMT, ALP
16 Consider the AST/ALT ratio AST/ALT > ……… severe liver damageAST/ALT < 1 ……… mild liver damage
17 Enzymes of Clinical Significance Source of blood elevationALTASTGMTALPACPCKAMSLPSCHShepatopathyMI, hepatopathyhepatopathy (alcohol, drugs)biliary tract diseases, bone diseasesprostatic cancerMI (CK-MB), muscle diseasespancreatitishepatopathy (alcohol, drugs) – decreased
18 Catalytic concentration of some enzymes Reference values (serum)ALTASTLDCKkat/lkat/lup to 7.5 kat/lup to 4 kat/lsee also the lab manual
19 Q. 7 What enzymes might appear in blood In mild hepatocellular damage b) In serious hepatocellular damage
20 A. 7 a) Mild hepatocellular damage: enzymes from cytoplasm and/or membrane are released into ECF – ALT, GMT, ALPb) Severe hepatocellular damage:enzymes from mitochondria are released into ECF – AST
21 Write equations of reactions catalyzed by: ALT AST LD
22 ALT Reactionalanine oxoglutarate plné názvy enzymů !!
23 ALT Reactionalanine oxoglutarate pyruvate + glutamate