Enzymes

Helpfulness: 0
Set Details Share
created 6 months ago by Pmborton
4 views
book cover
Clinical Chemistry
No Chapter
updated 6 months ago by Pmborton
show moreless
Page to share:
Embed this setcancel
COPY
code changes based on your size selection
Size:
X
Show:
1

enzyme

specific biologic proteins that catalyze biochemical reactions without altering equilibrium point of reaction or being consumed or changed in composition; changes composition of substance it acts on; reactions essential to biological functions

2

cofactor

non-protein molecule necessary for enzyme activity

3

activator

inorganic cofactor that increases the catalytic activity of an enzyme when it binds to a specific site

4

coenzyme

organic cofactor; diffusible, heat-stable substance of low molecular weight that, when combined with an inactive protein (apoenzyme), forms an active compound or a complete enzyme (holoenzyme) which may be necessary for activity

5

inhibitor

organic or inorganic compound that eliminates or reduces a reaction; may act directly on the enzyme

6

apoenzyme

protein portion of an enzyme without the cofactor necessary for catalysis

7

holoenzyme

apoenzyme + coenzyme; active compound or complete enzyme

8

substrate

compound on which the enzyme acts

9

activity

in vitro characterization of the amount of substrate that is reacted upon to produce a product over specified unit of time

10

isoenzyme

difference forms of an enzyme which catalyze the same reaction; genetic or nongenetic causes; differentiated by physical properties-electrophoretic mobility, solubility, and resistance to activation

11

oxidoreductases

catalyze oxidation-reduction reactions; dehydrogenases, oxidases (transfer oxygen), reductases (transfer hydrogen), and peroxidases

12

transferases

transfer of a biochemical group such as an amino-, carboxyl-, methyl-, or phosphoryl- group

13

hydrolases

catalyze the cleavage of C-O, C-N, C-C bonds; amylase, urease, pepsin

14

lysases

hydrolyze C-C, C-O, C-N bonds by elimination with the formation of a double bond or the addition of a group to a double bond

15

isomerases

induce structural changes-interconversion of geometrical, optical, or positional isomers

16

ligases

joins two molecules; involved in DNA, RNA, and protein synthesis

17

Jody

One of the greatest people I know

18

International Union of Biochemists

nomenclature with lengthy descriptive name for function and substrate

19

Enzyme Commission of IUB

name based upon 6 classes of enzymes, numbering system, recommended names listed

20

TABLE 13.1

...

21

active site

deep cleft in the enzyme where it sterically binds to substrate

22

activation energy

energy required to raise all molecules in 1 mole of a compound to the transition state at the peek of the energy barrier

23

enzymes

lower the activation energy needed for a reaction

24

lag phase

early time in an assay when mixing occurs and temperature and kinetic equilibria are being established

25

linear phase

time in an assay is producing a constant amount of product per unit of time

26

substrate depletion phase

time late in an enzyme assay when the substrate concentration is falling

27

substrate concentration

rate at which an enzymatic reaction proceeds and whether the forward or reverse reaction occurs depends on several reaction conditions

28

Michaelis and Menton

described the role of substrate concentration in formation of the enzyme-substrate complex

29

K(m)

the substrate concentration at which the rate of the reaction is half the maximum rate

30

first order kinetics

state occurring when the rate of an enzyme reaction is proportional to the concentration of the substrate

31

zero order kinetics

state occurring when rate of an enzyme is independent of the concentration of the substrate

32

Michaelis-Menten characteristics

equilibrium among all components, E, S, ES, and P

33

rate limiting step

formation of product and enzyme from ES complex; max velocity is fixed; reaction rate is function of enzyme as substrate is abundant

34

Km

should be constant for the reaction to take place; rule of M-Menten

35

low Km

high affinity of the enzyme for the substrate - low [s] is needed to half-saturate the enzyme

36

High Km

low affinity of enzyme for substrate - high [s] is needed to half-saturate the enzyme

37

enzyme concentration

causes increased [ES} and overall reaction rate when excess substrate is also available

38

pH

can alter structure or charge of enzymes impacting function

39

7.0-8.0

optimal pH range of enzymes

40

buffers

help control pH

41

temperature

proportional to reaction rate; important for all reagents

42

cofactors

nonproteins that must bind to an enzyme before the reaction can occur; metallic and nonmetallic activators; may be either essential or help enhance; alter enzyme binding site, link substrate to enzyme or coenzyme, and undergo oxidation or reduction

43

activators

inorganic cofactors; metallic such as Ca2+, Fe2+, Mg2+, Mn2+, Zn2+, and K+; nonmetallic Br- and Cl-

44

coenzymes

organic cofactors; nucleotide phosphates and vitamins; 2nd substrate for enzymatic reaction; called prosthetic group when bound tightly to enzyme; NAD-NADP

45

cofactor

included in excess in lab assay to not impact reaction

46

inhibition

can be reversible, irreversible, or done by antibodies; changes enzyme binding site so substrate cannot connect

47

competitive reaction

...

48

competitive inhibition

...

49

noncompetitive reaction

...

50

noncompetitive inhibition

...

51

uncompetitive

inhibitor combines with ES complex; rare, more common in two-substrate reactions

52

irreversible inhibition

covalent combination with enzyme; effect progresses with time

53

inhibition by antibodies

access of substrate molecules to active center by steric hindrance; conformational change induced in molecule

54

diagnostic enzymology

changes in the activity or mass in serum or plasma of enzymes that are predominantly intracellular and normally present in serum at low levels only

55

international unit

the quantity of enzyme that catalyzes the reaction of 1umol of substrate per minute

56

kinetic measurement

amount of change produced in a defined time interval

57

fixed time reaction

measured after reaction has stopped

58

creatine kinase (CK)

associated with ATP regeneration in contractile or transport systems; involved in storage of high-energy creatine phosphate; located in skeletal muscle, heart muscle, and brain tissue; significant in acute MI, muscular dystrophy, CVA, seizures, nerve degeneration, shock, hypothyroidism, malignant hyperplasia, Reye's syndrome; catalyzes both forward and the reverse reactions involving phosphorylation of creatine or ADP; hemolysis will elevate, inactivated by light

59

CK-MM - CK3

predominates in skeletal muscle (99%) and cardiac muscle (70%)

60

CK-MB - CK2

predominates in cardiac muscle (30%) and skeletal muscle (1%)

61

CK-BB - CK1

predominates in brain tissue; also found in bladder, lung, prostate, uterus, colon, stomach, and thyroid

62

reverse reaction

the more commonly used CK assay; 2-6x faster, less interference, pH 6.8

63

lactate dehydrogenase (LD)

found in the heart, liver, skeletal muscle, kidney, erythrocytes; 5 isoenzymes; significant in pernicious anemia, hemolytic disorders, viral hepatitis, cirrhosis, acute MI, pulmonary infarct, skeletal muscle disorders, and leukemia; catalyzes interconversion of lactic and pyruvic acids using coenzyme NAD in either forward or reverse direction; both used but reverse 3x faster but subject to substrate exhaustion and loss of linearity; hemolysis increases; unstable in any temperature, preferred storage 25C

64

LDH-1

located in heart and RBCs; MI and hemolytic anemia

65

LDH-2

located in heart and RBCs; megaloblastic anemia, acute renal infarct, hemolysis

66

LDH-3

located in lungs, lymphocytes, spleen, and pancreas; increased in pulmonary embolism, extensive pulmonary pneumonia, lymphocytosis, acute pancreatitis, and carcinoma

67

LDH-4

located in liver; increased in hepatic injury or inflammation

68

LDH-5

located in skeletal muscle; increased in skeletal muscle injury

69

aspartate aminotransferase (AST) (SGOT-serum glutamic-oxaloacetic transaminase)

transfers amine group between aspartate and alpha-keto acids which are ultimately used for energy; located in cardiac tissue, liver, skeletal muscle; small amount in RBCs, kidney, pancreas; significant in hepatocellular disorders (viral hepatitis, cirrhosis), skeletal muscle disorders (muscular dystrophy, inflammatory conditions), pulmonary embolism, AMI; very high in acute hepatocellular disorders

70

alanine aminotransferase (ALT) (SGPT-glutamic-pyruvic transaminase)

catalyzes transfer of amino group from alanine to alpha-ketoglutarate to form glutamate and pyruvate; distributed in many tissues with highest concentration in liver; significant in hepatic disorders-highest in hepatocellular than extra and intra hepatic disorders; high than AST in acute inflammatory conditions in the liver; coupled enzymatic reaction using lactate dehydrogenase as indicator enzyme, which catalyzes reduction of pyruvate to lactate with simultaneous oxidation of NADH; stable 3-4 days at 4C, relatively unaffected by hemolysis

71

alkaline phosphatase (ALP)

catalyzes hydrolysis of phosphomonoesters, nonspecific and reacts with many different substrates; found in intestine, liver, bone, spleen, placenta, kidney, and on cell surface of most human tissues; sinusoidal and bile canalicular membranes of liver, osteoblasts of bone; significant in hepatobiliary (biliary tract obstruction) and bone (Paget's disease, osteomalacia, rickets, hyperparathyroidism, osteogenic sarcoma) disorders; various methodologies-continuous-monitoring technique based on molar absorptivity of p-nitrophenol (Bowers and MaComb); isoenzymes-immunochemical methods; electrophoresis; unstable-hemolysis, run ASAP, ingestion of high-fat meal; increased in children during bone growth and development; functions at 9.0-10.0 pH

72

acid phosphatase (ACP)

hydrolase that catalyzes similar reactions to ALP; functions at 5.0 pH; found in prostate, bone, liver, spleen, kidney, erythrocytes, platelets, but prostate is the richest; significant in prostatic carcinoma, hyperplasia, and surgery, osteoclasts, Paget's, breast cancer with bone metastases, Gaucher's; PSA is best indicator for prostate cancer; forensic rape investigations; uses acidic pH p-nitrophenol; serum should be separated ASAP, used immediately and either frozen or acidified to <6.5 pH which makes it stable for 2 days at RT

73

y-Glutamyltransferase (GGT)

transfer of residue from peptides to amino acids, H2O2 and other small peptides; specific function not clear but involved in peptide and protein synthesis, transport of amino acids across cell membranes, regulation of tissue glutathione levels; found in kidney, brain, prostate, pancreas, liver (epithelial cells lining biliary ductiles) so very sensitive to liver damage and hepatocellular disorders; significant also in alcoholism, acute pancreatitis, diabetes mellitus, MI; normal in skeletal disorders and pregnancy; p-mitroanilide is most widely accepted substrate in analysis; residue is transferred to glycylglycine releasing p-nitroalanine; stable for 1 week at 4C, not affected by hemolysis

74

amylase (AMS)

hydrolase that catalyzes breakdown of starch and glycogen; requires calcium and chloride ions for activation; found in acinar cells of pancreas and salivary glands; lower concentration in skeletal muscle and small intestine; small so it appears in urine; significant in acute pancreatitis, disorders causing salivary gland lesions (mumps, parotitis), intraabdominal diseases; four main approaches for assays: amyloclastic, saccharogenic, chromogenic, and continuous monitoring; stable for 1 week at room temperature, 2 months at 4C; plasma triglycerides in acute pancreatitis with hyperlipemia; administration of morphine and other opiates

75

lipase (LPS)

hydrolyzes ester linkages of fats to produce alcohols and fatty acids; catalyzes partial hydrolysis of dietary triglycerides in the intestine with production of long chain fatty acids; primarily in pancreas; also in stomach and small intestine; significant in acute pancreatitis, other intraabdominal diseases (duodenal ulcers, perforated peptic ulcers, intestinal obstruction, acute cholecystitis); estimation of liberated fatty acids (triolein is substrate), turbidimetric methods, colorimetric methods; stable in serum for 1 week at RT and 3 weeks 4C; hemolysis causes falsely low levels-hemoglobin inhibits lipase

76

glucose-6-phosphate dehydrogenase (G-6-PD)

catalyzes the oxidation of glucose-6-phosohate to 6-phosphogluconate- 1st step in pentose-phosphate shunt of glucose metabolism for production of NADPH; found in adrenal cortex, spleen, thymus, lymph nodes, lactating mammary glands, erythrocytes; significant in G6PD deficiency (sex linked trait), drug induced hemolytic anemia, MI, and megaloblastic anemia; red cell hemolysate used to assay for deficiency using serum for elevation evaluation

77

macroenzymes

high molecular mass forms of the serum enzymes (ACP, ALP, ALT, amylase, AST, CK, GGT, LDH, lipase) that can be bound to either an immunoglobulin (macroenzyme type 1) or a nonimmunoglobulin substance (macroenzyme type 2); unexplained persistence increase of enzyme concentrations in serum; accumulate in plasma because their high molecular masses prevent them from being filtered out of plasma by the kidneys impacting assessment of increased enzyme levels-false increase; blocking of activity of bound enzyme-false decrease

78

drug metabolizing enzymes

transform xenobiotics into inactive, water-soluble compounds for excretion through kidneys; transform inactive prodrugs into active drugs, convert xenobiotics into toxic compounds, prolong elimination half-life; catalyze addition or removal of functional groups through hydroxylation, oxidation, dealkylation, dehydrogenation, reduction, deamination, desulfuration (phase I reactions)

79

P-450 (CYP 450)

mediates phase I reactions in >50% of drug metabolisms; genetic variants impact drug metabolism-fast and slow acetylators- pharmacogenetic testing is helpful to base medication on acetylator status

80

phase II reactions

transformed into more polar compound through conjugation