(13) FACTORS THAT INFLUENCE ENZYMATIC RXNs

Question 1

➢ The greater the substrate added = the ↑ the rate of enzymatic activity but when the addition of substrate conc. reaches the maximum value, the ↑ enzyme conc. already stops. = aka SATURATION KINETICS

Answer 1

Substrate Concentration

Question 2

• Used in lab
• Only a fixed # of substrate (in excess) is converted to product per second.

Answer 2

Zero-order

Question 3

• Rxn rate is proportional to substrate conc.

Answer 3

First-order

Question 4

Serial dilution is performed when enzyme result is extremely high that the machine cannot read the enzymatic activity.

Answer 4

• Serum is diluted then re-test.
• Do not report the result of the diluted serum.
• Compute (multiply the dilution factor to the result of diluted serum).

Question 5

➢ ↑enzyme conc. = faster rxn
➢ More enzyme present = more enzyme will bind to the available substrate

Answer 5

Enzyme Concentration

Question 6

➢ Hydrogen ion conc.
➢ Most physiologic rxn occur in range of 7.0-8.0 (near plasma pH)
➢ Extreme pH level (too ↓↑) leads to protein denaturation.

Answer 6

pH

Question 7

➢ Enzymes that can resist extreme pH levels:

Answer 7

Phosphatases such as ALP (9-10pH), ACP (3-5 pH)

Question 8

Optimum enzyme activity temp:

Answer 8

Body Temp

Question 9

2 temps for enzyme testing:

Answer 9

Room temp & Warm/Body temp

Question 10

Temperature

Answer 10

• Incubation time in room temp is longer.
• Incubation time in body temp is shorter.

Question 11

Incubation temp increase by 10°C → there is a characteristic 2-fold increase in the enzyme activity.

Answer 11

Temp. coefficient/Cube temp??:

Question 12

@low temp, enzymes ____

Answer 12

become inactivated (reversible)

Question 13

Start of denaturation process (irreversible)

Answer 13

> 37°C

Question 14

Significant enzyme denaturation is seen at this temp

Answer 14

56°C

Question 15

Enzymes are completely inactivated

Answer 15

60-65°C:

Question 16

Most enzymes are thermolabile (inactive @ high temp)

Answer 16

• Bcos protein part of enzyme would cause denaturation rxn @ very ↑temp.
• Ex. ALP

Question 17

NOTE:

Answer 17

➢ Placental ALP (specifically regan ALP) is thermos-stable.
➢ Assay temp should be constant ±0.1°C @ which the enzyme is active @ 25°C, 30°C, 37°C
➢ Multiple freeze-thawing of enzymes leads to protein denaturation making it inactive (should only be performed once).

Question 18

-20°C or colder (freezer temp)

Answer 18

SERUM

Question 19

stored at Ref temp

Answer 19

SUBSTRATE & COENZYME

Question 20

Room temp

Answer 20

ENZYMES W/ LDH

Question 21

➢ Can enhance enzyme activity

Answer 21

Cofactors

Question 22

Activators:

Answer 22

*Metallic: Ca2+, Fe2+, Mg2+, Mn2+, Zn2+
*Non-metals: Br, Cl

Question 22

Activators:

Answer 22

*Metallic: Ca2+, Fe2+, Mg2+, Mn2+, Zn2+
*Non-metals: Br, Cl

Question 23

*Serve as second substrates for enzymatic rxns (NAD)

Answer 23

Coenzymes (prosthetic groups)

Question 24

Interferes w/ enzymatic rxn w/ the substrate

Answer 24

Inhibitors

Question 25

can physically bind to the active site of the enzyme (can compete to the substrate)

Answer 25

Competitive inhibitor

Question 26

o binds to the enzyme in a place other than the active site
o Binds to allosteric site

Answer 26

Non-competitive

Question 27

o binds w/ active site of enzyme.
o Binds to enzyme-substrate complex

Answer 27

Uncompetitive

Question 28

➢ RBCs have enzymes inside,
➢ this causes false ↑ in enzyme conc.

Answer 28

Hemolysis

Question 29

➢ Lactescent → False ↓ in enzyme conc.

Answer 29

Lipemic/milky