ENZYMES IN MEDICINE
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ENZYMES IN MEDICINEENZYMES IN MEDICINE
• 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
ENZYMES IN CLINICAL DIAGNOSIS
secretory - produced by tissues (namely liver), acting in plasma – prothrombin, plasminogen, cerruloplasmin, choline esterase; lipoprotein lipase
Enzymes
intracellular – function intracellulary, have no physiological use in 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
• 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
(determined in clinical laboratory – each lab 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
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
Examples of enzymes commonly assayed for diagnostic purposes
Enzyme Location Cause of elevated plasma level
Acid phosphatase - ACP Prostate Prostatic cancer
Alkaline phosphatase – ALP Bone, liver Rickets, hypoparathyroidism, osteomalacia, obstructive jaundice, cancer of bone/liver
Alanine aminotransferase – ALT Liver (muscle, Hepatitis, jaundice, circulatory heart, kidney) faillure with liver congestion
Aspartate aminotransferase – AST Heart, muscle, Myocardial infarction, muscle red cells, liver damage, anemia, hepatitis,
circulatory faillure with liver congestion
Amylase - AM Pancres Acute pancreatitis, peptic ulcer
-Glutamyl transferase – GMT Liver, kidney, Hepatitis, alcoholic liver pancreas damage, cholestasis
Examples of isoenzymes commonly assayed for diagnostic purposes
Enzyme Location Cause of elevated plasma level
Creatine kinase – CKCK-MB Heart Myocardial infarctionCK-MM Skeletal muscle Muscular dystrophy
Lactate dehydrogenase – LDLD1 > LD2 Heart, kidney, Myocardial infarction, kidney blood cells disease, megaloblastic anemia,
leukemia
LD2, LD3 LeukemiaLD5 Liver, muscle Liver disease, muscle damage
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
ENZYMOTHERAPYOrally 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,
chymotrypsin, papain (Carica papaya), bromelain (ananas) = combination of enzymes with different specificity, pH optimum, stability, interaction with inhibitors and antiproteinases
multiple action
• 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 –
1-antitrypsin, 2-macroglobulin complexes
direct proteolytic action, degradation of adhesive
molecules, secretion of cytokins (tranforming growth factor TGF-), modulation of receptor function
not fully clarified
?
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
NUCLEOTIDESNUCLEOTIDESStructure, FunctionStructure, Function
NUCLEOTIDE STRUCTURENUCLEOTIDE STRUCTURE
Nucleotides
nitrogenous base + pentose + phosphate group(s)
purine ribose 1 - 3
pyrimidine deoxyribose
other (nicotinamide)
Nucleosides
NC
C
CCN
N
NH
CH
NH2
|
H
NC
C
CCN
N
NH
CH
O||
H
H2N
NC
C
CCNH
O
H
H
O|| CH3
adenineadenine guanineguanine
cytosinecytosine thyminethymine uraciluracil
THE NITROGENOUS BASESTHE NITROGENOUS BASESTHE NITROGENOUS BASESTHE NITROGENOUS BASES
NC
C
CCNH
O
NH2
|
H
H NC
C
CCNH
O
O||
HH
H
Purine bases
Pyrimidine bases
NUCLEOSIDENUCLEOSIDENUCLEOSIDENUCLEOSIDE
• A sugar - base combination.
OHOCH2
H HHH
OH H
BaseBase
SugarIn this case deoxyribose
SugarIn this case deoxyribose
-N-glycosidiclinkage
-N-glycosidiclinkage
N
O
H HHH
OH H
HOCH2 O
H HHH
OH OH
HOCH2
O
H HHH
OH
HOCH2
NC
C
CCNO
NH2
|
H
H
NC
C
CCNO
H
H
CH3
O
NC
C
CCNO
H
H
O
H
OH
thymine uracil
cytosine
deoxythymidine uridine
cytidine
O
H HHH
OH H
O
H HHH
OH
O
H HHH
OH
HOCH2
HOCH2
HOCH2
OH
OH
NC
C
CCN
N
N
CH
NH2
|
H
NC
C
CCN
N
N
CH
OH|
H
NC
C
CCN
N
N
CH
O
H
H2N
adenine guanine
hypoxanthine
deoxyadenosine guanosine
inosine
!
NUCLEOTIDESNUCLEOTIDESNUCLEOTIDESNUCLEOTIDES
5’-OH on the sugar of a nucleoside is converted into a phosphate ester.
NC
C
CCN
N
N
CH
NH2
|
H
O-O-P-O-CH2
H HHH
OH H
|
O- -
O||
deoxyadenosine monophosphate(dAMP)
deoxyadenosine monophosphate(dAMP)
Each is named based on sugar and base nameand then the number ofphosphates is indicated.
adenine
phosphate chain
ribose
CH2 O
OH OH
N
NN
N
NH2
OPOPOPO-
O
O-
O
O-
O
O-
ATPATP - - adenosine triphosphateadenosine triphosphate
ATPADP
AMP
NUCLEOTIDE FUNCTIONNUCLEOTIDE 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
NAD+NAD+
OCH2
OH OH
OPO
O-
O
OCH2
OH OH
OPO
O-N
NN
N
NH2
N+
C
O
NH2
reactivesite
nicotinamide
adenine
ribose
OCH2
OH OH
OPO
O-N
NN
N
NH2
O
C HH
CH OH
CH OH
CH OH
CH H
N
NH3C
H3C N
NH
O
O
ribose
adenine
riboflavin
reactive site
FAD FAD
Coenzyme ACoenzyme A
phosphorylated ADP
pantothenateunit
CH2 O
O OH
N
NN
N
NH2
OPOPO
O-
O
O-
P O-O
O-
C-CH2-CH2-N-C-C-C-CH2
OO H
CH3HO
CH3
HH-N
CH2-CH2
SSulfhydryl
group
O
H
H
OH
O H
OHOH
H
OHH
OH
CH2OH
H -D-glucose -D-glucose
OH
OH
H
H
H
OH
CH 2 OH
H
OH
OH OH
H
H
H
OH
CH 2 OH
H
OH
O
Non-polar tail
O CO
O CO
C
C
CH2
H
H
H
PO
O
O-
2-O-
cAMP
cAMP – cyclic adenosine monophosphate
NC
C
CCN
N
N
CH
NH2
|
H
O
H HHH
H
-
P O
O
O
O-
O
CH2
cAMP – cyclic adenosine monophosphate
-intracellular mediator, second messenger of hormonal signal tranduction via adenylate cyclase cascade- mechanism of action: allosteric effector
CH2-O-C
HO HOHO
C-O-CH
CH2-O - P
O
OO
SCoA
HO
O
SCoA
HOHO
P~
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