Enzymes + Protein Function + Protein Analysis

Question 1

what’s the difference between cofactors and coenzymes?

Answer 1

cofactors are inorganic molecules or metal ions

coenzyme are organic molecules.

Question 2

how can one increase Vmax?

Answer 2

increase amount of enzyme

Question 3

what’s the michaelis-menten equation?

Answer 3

v = (Vmax[S])/(Km + [S])

Question 4

What does Hill’s Coefficient measure?

Answer 4

it quantifies an enzyme’s cooperativity

Question 5

When Hill’s coefficient is greater than 1, what is indicated?

Answer 5

> 1: positively cooperative binding

after one substrate binds, it’s easier for other substrates to bind (enzyme affinity for other substrates increases)

Question 6

when hill’s coefficient is less than 1, what is indicated?

Answer 6

<1: negatively cooperative binding

after one substrate binds, it’s harder for other substrates to bind (enzyme affinity for other substrates decreases)

Question 7

when hill’s coefficient is 1, what is indicated?

Answer 7

the enzyme does not have cooperative binding

Question 8

what is the shape of the michaelis-menten plot when an enzyme exhibits cooperative binding?

Answer 8

sigmoidal curve (S curve)

Question 9

What’s the relationship between enzyme activity and temperature?

Answer 9

enzyme activity doubles for a 10 degree increases in temperature until optimum temperature
after optimum temperature, enzyme activity quickly drops off due to denaturation

Question 10

what is the optimal pH of pepsin, an enzyme that functions in the stomach?

Answer 10

pH 2

Question 11

what is the optimal pH for pancreatic enzymes that function in the small intestine?

Answer 11

pH 8.5

Question 12

what’s the effect of competitive inhibition on Km and Vmax?

Answer 12

increases Km

no effect on Vmax

Question 13

what’s the effect of noncompetitive inhibition on Km and Vmax? what’s the affinity of a noncompetitive inhibitor to enzyme and enzyme-substrate complex?

Answer 13

noncompetitive inhibitors bind equally well to enzyme and enzyme-substrate complex
no effect on Km
decreases Vmax

Question 14

what’s the effect of uncompetitive inhibition on Km and Vmax? what do uncompetitive inhibitors bind?

Answer 14

uncompetitive inhibitors only bind enzyme-substrate complex
decrease Km
decrease Vmax

Question 15

what’s the effect of mixed inhibitors on Km and Vmax?

Answer 15

decrease Vmax
increase Km if preferentially bind enzyme over enzyme-substrate complex
decrease Km if preferentially bind enzyme-substrate complex over enzyme

Question 16

what is the structure and function of collagen?

Answer 16

trihelical structure

makes up ECM in connective tissue

Question 17

what is the function of elastin?

Answer 17

restoring shape of tissue

part of ECM and it stretches and recoils like a string

Question 18

what is the function of keratin and what type of filament is it?

Answer 18

keratin is an intermediate filament

provides mechanical integrity to cells (nails, hair)

Question 19

what is the function of actin?

Answer 19

microfilaments involves in muscle contraction and cytokinesis (cleavage furrow)

Question 20

what is the function of tubulin?

Answer 20

chromosome separation and intracellular transport

Question 21

where is the negative and positive end of tubulin located in a cell?

Answer 21

negative end is close to nucleus and positive end is near cell membrane

Question 22

in what direction does kinesin travel on microtubules?

Answer 22

travels from negative end towards positive end

Question 23

in what direction does dynein travel on microtubules?

Answer 23

travels from positive end to negative end

Question 24

does kinesin or dynein bring vesicles of neurotransmitters to the synaptic terminal?

Answer 24

kinesin - travels from negative end of microtubule to positive end of microtubule (tubulin has negative end close to nucleus, soma, and positive end close to synaptic terminal)

Question 25

what is the function of cadherins?

Answer 25

glycoprotein that connects similar cells together in a calcium dependent manner

Question 26

what is the function of integrins?

Answer 26

binds and communicates with ECM; and cellular signaling

Question 27

what is the function of selectins?

Answer 27

bind carbohydrates on other cells; important for immune system

Question 28

what is opsonization? (hint: antibody)

Answer 28

when an antibody marks a pathogen for destruction by white blood cells

Question 29

what is agglutinating? (hint: antibody)

Answer 29

when an antibody binds an antigen and leads to clumping of insoluble complexes

Question 30

in anion exchange chromatography, what is the charge of the resin?

Answer 30

in anion exchange chromatography, the resin is positively charged

Question 31

in size exclusion chromatography, do larger or smaller compounds elute earlier?

Answer 31

larger compounds elute earlier since smaller compounds get stuck in pores

Question 32

what is Edman degradation?

Answer 32

sequencing proteins of 50-70 amino acids

removes N-terminal amino acid and identifies it using mass spec, and repeat

Question 33

what is the Bradford protein assay?

Answer 33

method of determining protein concentration

1. known [protein] + comassie blue –> absorbance –> standard curve
2. unknown [protein] + coomassie blue –> absorbance –> determine [protein]

Question 34

in the Bradford protein assay, does a more blue color indicate less or more protein concentration?

Answer 34

for bradford protein assay, more blue = higher [protein]

Coomassie blue is green-brown and turns blue when mixed with protein