Enzymes

Question 1

Enzyme (mechanism of action)

Answer 1

• catalysts (not consumed in rxns)
• decrease activation energy needed to form product for more rapid formation
• S + E –> ES –> P + E

Question 2

Factors affecting enzyme reactions

Answer 2

• time: fixed or kinetic
• substrate level
• pH of mixture (pH 7-8)
• temperature (30-37C)
• enzyme concentration
• activators/inhibitors

Question 3

competitive inhibitors

Answer 3

bind to active site of enzyme

Question 4

uncompetitive inhibitors

Answer 4

bind to ES complex

Question 5

non-competitive inhibitors

Answer 5

bind to another site (not active site) on enzyme

Question 6

Enzyme classes

Answer 6

• oxidoreductases
• transferases
• hydrolases
• lyases
• isomerases
• ligases

Question 7

Enzyme unites of measure

Answer 7

• international unit (amt of enzyme which can convert 1 umole of substrate to product per min)
• systemic unit (amt of enzyme that catalyzes 1 mole of substrate per second = katal unit)

Question 8

isoenzymes

Answer 8

• forms of enzymes that are only structurally different
• can catalyze same reaction as enzyme
• separated by electrophoresis or Ag-Ab rxn

Question 9

Creatine kinase

Answer 9

• associated with ATP regeneration in muscle and transport systems
• found in heart, muscle, brain tissue
• elevations associated with AMI, muscular dystrophy, CNS seizures/shock, hypothyroidism, Reye’s syndrome, malignant hyperpyrexia

Question 10

CK methods

Answer 10

• forward rxn: measure absorbance

- reverse rxn: commonly used method

Question 11

CK isoenzymes

Answer 11

• M and B units of dimer
• CK1 or BB isoenzyme (brain)
• CK2 or MB isoenzyme (heart muscle)
• CK3 or MM isoenzyme (skeletal muscle)

Question 12

Aspartate Aminotransferase (AST)

Answer 12

• transfers amino group to make new AA
• pyridoxal phosphate is coenzyme
• found in cardiac, liver and skeletal tissue
• significant in viral hepatitis

Question 13

AST methods

Answer 13

• amino acid transferase rxn with alpha keto acid oxoglutarate
• read NAD absorbance

Question 14

LD method

Answer 14

• uses pyruvate to lactate
• NAD absorbance is read
• increases of LD only if cell damage (because its a cytoplasmic enzyme)

Question 15

LD isoenzymes

Answer 15

• uses H and M subunits in tetramers
• 5 possible isoenzymes
• LD1 > LD2 in heart attack
• LD4 and LD5 increase in liver disease
• LD3 is seen in pulmonary disorders
• LD6 is alcohol dehydrogenase

Question 16

Lactate Dehydrogenase

Answer 16

• In AMI: rise, peak, remain elevated for 10 days

- in hepatic disorders: LD5 increase

Question 17

Heart attack tests

Answer 17

CK, AST, LD and troponin

Question 18

Liver disease tests

Answer 18

AST, LD, ALT, gamma-GT, ALP

Question 19

Alanine Transferase (ALT)

Answer 19

• found in liver (liver specificity)
• increases in viral hepatitis, so does AST
• in cirrhosis AST > ALT

Question 20

ALT method

Answer 20

• rxn is AA transfer from alaning to keto acid oxoglutarate

- read NAD absorbance

Question 21

Alkaline Phosphatase (ALP)

Answer 21

• hydrolyzes phosphate groups from molecules at alkaline pH
• Mg is activator
• found in most cell membranes
• found in intestine, kidney, bone, liver and placenta
• increases associated with liver or bone

Question 22

ALP method

Answer 22

• ALP is hydrolase enzyme

- method uses para-nitro-phenol phosphate (PNPP)

Question 23

ALP isoenzymes

Answer 23

• 5 isoenzyme forms
• liver and bone overlaps on electrophoresis
• do immunochemical method for Bone ALP
• inhibition testing uses phenylalanine

Question 24

Gamma-GT (Gamma glutaryl transferase)

Answer 24

• involved in the transfer of gamma-glutamyl group to other AA
• found in all cells but muscle
• originates in hepatobiliary system
• can increases with alcohol and cirrhosis

Question 25

Amylase

Answer 25

- found in salivary glands and in pancreas - 3 isoenzymes for salivary amylase, 3 for pancreatic amylase - hydrolyzes alpha-1,4-glycosidic bonds - increase seen in pancreatitis, mumps

Question 26

Amylase methods

Answer 26

- Classic iodometric method - Saccharogenic method - Enzymatic method - read absorbance

Question 27

Classic iodometric method (amylase)

Answer 27

- iodine used to react with starch substrate | - when starch is all glucose, no iodine rxn

Question 28

Saccharogenic method (amylase)

Answer 28

measured glucose formed by amylase action on starch substrate

Question 29

Enzymatic method (amylase)

Answer 29

uses artificial substrate and measures glucose

Question 30

Lipase

Answer 30

- pancreatic enzyme - used in conjunction with amylase to identify pancreatitis - method uses artificial substrate of triolein and links rxns to dye color change