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29 Cards in this Set
- Front
- Back
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characteristics of an enzyme |
enzymes are catalysts that speed up reactions by lowering the amount of activation energy necessary for the reaction to occur majority are large proteins can be denatured by heat or shaking which will disrupt the tertiary structure of the enzyme. always end in "ase" |
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Activity assay |
measure the actual enzyme activity upon a substrate; activity correlates to concentration. |
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Mass assays |
measure actual concentration; typically done by immunoassay using monoclonal antibodies. |
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mechanisms for increase concentrations in enzymes |
tissue necrosis increased cell membrane permeability increased synthesis (tumors) obstructive disease decreased catabolism |
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decreased concentrations of enzymes |
decreased synthesis ( genetic, nutritional, inhibitory) excretion (renal disease) |
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Factors that influence enzyme reactions |
Substrate concentration enzyme concentration pH temp (37C) Cofactors (Mg,Fe,Zn) Inhibitors (NaF) |
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Substrate concentration, first order reaction |
if substrate concentration is low then there will be a direct proportion between the reaction rate and the amount of substrae concentration. |
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substrate concentration, zero order reaction |
if the substrate concentration is in great excess compared to the enzyme concentration, the enzyme binding sites will become saturated. at this point the reaction velocity becomes independent of the substrate concentration. this is called zero order kinetics as at maximum velocity the rate of increase in the speed of the reaction falls to zero. |
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temp for enzyme assay |
too high denaturization may occur, too low enzymes become reversibly inactive |
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Cofactors |
Mg, Fe, Zn necessary for reaction to occur, may be second substrate |
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inhibitors |
if present will inhibit enzyme action |
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when a reaction is performed in zero order kinetics. |
the rate of the reaction is independent of the substrate concentration |
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activity of enzymes in serum may be determined rather than concentration because |
the amount of enzyme is too low to measure. |
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end point monitoring methods |
measure the end point when the reaction is done |
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multipoint monitoring methods |
measures the change up to the end point. |
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most common cause of non-linearity in the enzymatic assay |
all substrate used up early in the reaction time. For the remainder of the reaction, the rate change is minimal with the implication that the coenzyme concentration is very low. |
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coupled enzyme assays |
NADH absorbs at 340 and 366nm, NAD does not. Can either have a reaction where NAD+ is reduced to form NADH+ and the absorbance will increase at 340 nm. or have a reaction where NADH is oxidized to form NAD+ and the absorbance at 340 nm will decrease |
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physiologic enzyme origins |
skeletal tissue cardiac tissue liver bone pancreas intestine prostate blood |
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Creatin Kinase CK |
Dimer - 2 polypeptide chains, M and B isoenzyme fractions: MM, MB, BB MM: mainly found in skeletal muscle MB: found in cardiac and some skeletal muscle. BB: found in brain tissue. CK-MB used in the assessment of myocardial infarction; analyzed by mass assay Total CK is used to evaluate muscular disorders like muscular dystrophy |
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Lactate Dehydrogenase LD |
Tetramer - 4 polypeptide chains with subunit coposition of M and H 5 isoenzyme fractions: LD4,5: liver; sensitive to temperatures lower than room temp. LD3: may be elevated in some leukemias and lymphomas. LD (total) is found in extremely high concentration in RBC's; therefore, analysis greatly affected by hemolysis |
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Aspartate Aminotransferase AST |
not very specific elevated in hepatocellular disorders (hepatitis, cirrhosis) affected by hemolysis used to be called SGOT |
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Alanine Aminotransferase ALT |
found mainly in liver more specific than AST and LD elevated in hepatocellular disorders used to be called SGPT |
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Alkaline phosphatase ALP |
found in the following tissue sources: liver, bone, intestine, kidney and placenta children and women in their 3rd trimester of pregnancy have higher levels elevated in: obstructive liver disease, Paget's disease, metastases to bone, liver, pancreas; Regan isoenzyme fraction present in small percentage (3-15%) of cancer patients, blockage in urobilliary system. |
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Gamma glutamyltransferase |
present in hepatocytes elevated in obstructive disorders; more sensitive than ALP. sensitive indicator of alcoholic liver disease. |
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Acid phosphatase ACP |
found in red cells and prostate elevated in prostatic carcinoma may be used in rape cases (found in seminal fluid), vaginal washings examined. |
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Amylase AMY |
found in saliva and pancreatic tissue elevated in pancreatitis serum and urine measurements done: urine levels may be higher and persist longer since amylase is rapidly filtered from the blood into the urine. |
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Lipase LPS |
found in pancreatic tissue elevated in pancreatitis more sensitive and specific than amylase; elevations are higher and persist for longer period of time breaks down lipids |
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Acetylcholinesterase AChE |
true, tissue, or RBC cholinesterase found in nerve tissue, RBS, lung, spleen, and grey matter of brain. important role in regulation of nerve transmission - hydrolyzes acetylcholine we dont usually measure this |
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Serum Cholinesterase SChE |
called pseudocholinesterase found in serum, liver, pancreas, heart and other tissues hydrolyzes acetylcholine more rapidly than AChE inhibited by organophosphates (found in pesticides, herbicides, fungicides,) patients who are poisoned by organophosphates will have nerve weakness, may have trouble breathing or walking. |
