Medicinal Enzyme and Nucleotide


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Basic study of enzyme for medicine study.

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Medicinal Enzyme and Nucleotide

  2. 2. • Diagnostic indicators – the activities of many enzymes are routinely determined in plasma ( rarely in tissue biopsies) for diagnostic purposes in diseases of the heart, liver, skeletal muscle, pancreas and other tissues - enzyme diagnostics • Therapeutic agents – several enzymes are used as drugs; new approach - enzymotherapy • Diagnostic tools – use as chemicals in clinical laboratory assays
  3. 3. ENZYMES IN CLINICAL DIAGNOSIS plasma – secretory - produced by tissues (namely choline Enzymes use in liver), acting in prothrombin, plasminogen, cerruloplasmin, esterase; lipoprotein lipase intracellular – function intracellulary, have no physiological plasma - membrane bound – ALP, GMT - cytosolic – ALT, AST, LD, MDH - mitochondrial – AST, GMDH - lysosomal ACP - tissue specific – glucose-6-phosphatase – liver amylase – pancrease LD1 – heart
  4. 4. • Healthy individuals - levels of intracellular enzymes fairly constant, low – the rate of enzyme release from damaged cells into plasma balanced by the rate of removal of enzyme protein from plasma ⇓ Physiological enzyme levels ⇒ reference values of the enzyme activities lab (determined in clinical laboratory – each has its own reference values) • Elevated enzyme activity in the plasma – reflect tissue damage accompanied by increased release of intracellular enzyme Skeletal muscle during exertion – physiologically elevated levels of muscle enzymes in plasma • Many diagnostically important enzymes = isoenzymes – pattern of isoenzymes in plasma (determined electroforetically) – a means of identifying the damaged tissue
  5. 5. ALTERATION OF ENZYME PLASMA LEVELS Increased values – increased cell membrane permeability anoxia, disturbances of energy metabolism ⇒ cytosolic enzymes – ALT, LD, CK - cell necrosis ⇒ membrane-bound enzymes – ALP, GMT mitochondrial enzymes – AST, GMDH - induction of the enzyme synthesis ← drugs – ALP, GMT Decreased values – inhibition of the activity ← drugs - inhibition of the synthesis ← cell damage, drugs
  6. 6. Examples of enzymes commonly assayed for diagnostic purposes Enzyme level Location Cause of elevated plasma Acid phosphatase - ACP Prostate Prostatic cancer Alkaline phosphatase – ALP hypoparathyroidism, Bone, liver Rickets, osteomalacia, obstructive jaundice, cancer of bone/liver Alanine aminotransferase – ALT circulatory Liver (muscle, heart, kidney) Aspartate aminotransferase – AST Heart, muscle, muscle red cells, liver liver Amylase - AM ulcer γ-Glutamyl transferase – GMT Hepatitis, jaundice, faillure with liver congestion Myocardial infarction, damage, anemia, hepatitis, circulatory faillure with congestion Pancres Liver, kidney, pancreas Acute pancreatitis, peptic Hepatitis, alcoholic liver damage, cholestasis
  7. 7. Examples of isoenzymes commonly assayed for diagnostic purposes Enzyme plasma level Creatine kinase – CK CK-MB CK-MM Lactate dehydrogenase – LD LD1 > LD2 kidney Location Cause of elevated Heart Skeletal muscle Myocardial infarction Muscular dystrophy Heart, kidney, Myocardial infarction, blood cells disease, megaloblastic anemia, leukemia LD2, LD3 LD5 damage Leukemia Liver, muscle Liver disease, muscle
  8. 8. ENZYMES IN THERAPY • Substitution of missing production of digestive enzymes – digestive enzymes – pepsin trypsin… • Removal of deposits of death tissue or fibrin (e.g. in lungs, eyes), treatment of skin defects – proteinases, nucleases, collagenase • Acceleration of fibrinolysis in lungs embolization (activation of plasmin and plasminogen) – streptokinase, urokinase
  9. 9. ENZYMOTHERAPY Orally administered enzymes – treatment of a variety disorders - digestive, gastrointestinal, pancreatic - inflammatory diseases, edema - immune and autoimmune diseases (arthritis, multiple sclerosis) - viral diseases (herpes, AIDS) - cancer Mixtures of enzymes of plant and/or animal origin - proteinases, amylase, lipase - administered as acidoresistent tablets • Pancreatin – trypsin, chymotrypsin, lipase, amylase • Wobenzym – pancreatic and plant proteolytic enzymes – trypsin, = optimum, chymotrypsin, papain (Carica papaya), bromelain (ananas) combination of enzymes with different specificity, pH stability, interaction with inhibitors and antiproteinases ⇒ multiple action
  10. 10. • Mechanism of resorption (transport of large macromolecules across the intestinal barrier) – paracellular transport, receptor mediated endocytosis and transcytosis • Mechanism of action – interaction with plasma antiproteinases – →complexes ? factor α1-antitrypsin, α2-macroglobulin direct proteolytic action, degradation of adhesive molecules, secretion of cytokins (tranforming growth TGF-β), modulation of receptor function not fully clarified
  11. 11. ENZYMES - USE IN LABORATORY ASSAYS Enzymes isolated from different sources - used for determination of various substances in the blood, plasma/serum and urine ← enzyme methods much more specific than chemical methods, the presence of relative substances with similar chemical properties does not hinder Components of commercial kits or diagnostic strips - determination of glucose - glucose oxidase, peroxidase cholesterol - cholesterol esterase, cholesterol oxidase peroxidase, urea – urease, ……. in blood, plasma, serum - proof of glucose (glucose oxidase), …….. in blood or urine (strips) Markes in the immunochemical analysis - ELISA (=enzyme-linked immunoadsorbent assay) – peroxidase, alkaline phosphatase
  13. 13. NUCLEOTIDE STRUCTURE Nucleotides nitrogenous base + pentose + group(s) purine ribose pyrimidine deoxyribose other (nicotinamide) Nucleosides phosphate 1-3
  14. 14. THE NITROGENOUS BASES Purine bases NH 2 | C N O || C adenine adenine N C HN CH HC C N C CH C N H N guanine guanine H 2N C N H N Pyrimidine bases O || C NH 2 | C N O C CH N H cytosine cytosine CH 3 N H O C C CH O || C CH N H thymine thymine HN CH C C H O N H uracil uracil
  15. 15. NUCLEOSIDE • A sugar - base combination. Base Base N O HOCH 2 H Sugar Sugar In this case In this case deoxyribose deoxyribose H H OH H H β-N-glycosidic β-N-glycosidic linkage linkage
  16. 16. thymine O O C C H HN CH N O OH H H H N C O H H OH CH N O H H OH OH CH N NH 2 | C CH deoxythymidine H C OH H C O HOCH 2 HOCH 2 H C O HOCH 2 H O CH 3 C HN uracil H cytidine cytosine H uridine
  17. 17. adenine NH 2 | C N C O N C HN CH HC C N H C H N H 2N C C N OH H H deoxyadenosine OH | C N C N H H OH H OH hypoxanthine CH HC C N ! O HOCH 2 H N H H OH OH H inosine N O HOCH 2 H N CH O HOCH 2 guanine H guanosine
  18. 18. NUCLEOTIDES 5’-OH on the sugar of a nucleoside is converted into a phosphate ester. NH 2 | C deoxyadenosine monophosphate deoxyadenosine monophosphate (dAMP) (dAMP) N Each is named based on sugar and base name and then the number of phosphates is indicated. C N CH HC O || - O-P-O-CH 2 | O- - H C N N O H H OH H H
  19. 19. ATP - adenosine triphosphate adenine phosphate chain O - O P O- O O P O- NH2 O O N P O O- CH2 N O OH ADP ATP N AMP ribose OH N
  20. 20. NUCLEOTIDE FUNCTION • Precursors of DNA, RNA - NTPs • Energy transport - ATP • Allosteric effectors of enzymes – ATP, ADP, AMP • Covalent modification of enzymes – ATP • Intracellular mediators (= second messengers) – cAMP, cGMP • Coenzymes – NAD+, NADP+, FAD, CoA-SH • Activated precursors of polysaccharaides, glycoproteins, proteoglycans, phospholipids, glycolipids – UDPG, UDPGA, UDPGal…, CDP-choline, CDP-diacylglycerol… • Active groups (group transport) – SAM, PAPS
  21. 21. NAD+ reactiv e site O O P O C O CH2 O O OH P O O - ribose CH2 nicotinamide N+ NH2 OH N O NH2 N O OH OH N N adenine
  22. 22. FAD O H3C N H3C reactive site NH N N H C H H C OH H C OH H C OH H C O H riboflavin NH2 N O O P O O - ribose CH2 N O OH OH N N adenine
  23. 23. Coenzyme A phosphorylated ADP pantothenate unit O H CH 3 O C-CH 2 -CH 2 -N-C-C-C-CH 2 H HO CH 3 H-N NH2 O O O N P O P O O- O- CH2 O N CH 2 -CH 2 S H N Sulfhydry l group O O P OO- OH N
  24. 24. CH2 OH CH2 OH H H OH O H OH H OH α -D-glucose H H O H OH O OH H H H CH2OH OH H β -D-glucose O H OH H H OH OH H OH
  25. 25. H O H C O C O H C O C O 2 O P -O- CH 2 - O Non-polar tail
  26. 26. cAMP – cyclic adenosine monophosphate cAMP
  27. 27. NH 2 | C N C N CH HC C N O CH 2 - O O P H H O N H H O H O- cAMP – cyclic adenosine monophosphate -intracellular mediator, second messenger of hormonal signal tranduction via adenylate cyclase cascade - mechanism of action: allosteric effector
  28. 28. O O O O SCoA C-O-CH SCoA HO HO HO HO CH2-O-C CH2-O - P ~ P HO HO