Enzymes 2

What is the Lineweaver-Burke equation?

1/v = (Km/Vmax)*(1/[S]} + (1/Vmax)

When substrate is low, what is the rate?

1st order

When substrate is high, what is the rate?

zero order

What is the upper limit for catalytic efficiency?

diffusion limit

What is Vmax?

theoretical max rate of rxn

Is Vmax ever achieved in reality?

No, requires infinite substrate

What is the x-axis of LWB plot?

1/[S]

What is the y-axis of LWB plot?

1/v

What is the slope of LWB plot?

Km/Vmax

What is the y-intercept of LWB plot?

1/Vmax

What is the x-intercept of LWB plot?

-1/Km

What happens to Km in Competitive Inhibition?

increases, lower affinity

Why does Km change in Competitive Inhibition?

more binding sites occupied, harder to bind substrate

What happens to Vmax in Competitive Inhibition?

no change

Why does Vmax not change in Competitive Inhibition?

inhibitor binds reversibly, can overcome by increasing substrate

What happens to the slope in Competitive Inhibition?

increases

What happens to the LWB plot in Competitive Inhibition?

same y-int, x-int moves towards zero

What is Ki?

inhibition constant

What does Ki measure?

affinity of an inhibitor for the enzyme

What does α (alpha) tell you?

how much the inhibitor changes the enzyme kinetics

What is α (alpha)?

1 + [I]/Ki

What does a high alpha mean?

stronger inhibitory effect

Why do we use α?

Instead of rewriting the MM equation every time, we multiply certain terms by α

What do we multiple alpha by in Competitive Inhibition?

Km

How do we calculate alpha for Competitive Inhibition?

slope₀ / slope_i

What is slope_i ?

a*Km/Vmax

What is the equation for Ki?

[I]/(m₀ / m_i - 1)

Why does Mixed Inhibition have TWO binding constants?

inhibitor can bind to both: Free enzyme (E) and Enzyme-substrate complex (ES)

What do Pure Noncompetitive Inhibitors bind to?

enzyme OR enzyme-substrate complex

What type of inhibitors are Irreversible Inhibitors seen as?

Pure Noncompetitive

Why types of bonds to reversible inhibitors bind to enzyme with?

noncovalent

Why types of bonds to irreversible inhibitors bind to enzyme with?

covalent

Where does a Pure Noncompetitive Inhibitor bind?

allosteric site

What’s the difference between Pure Noncompetitive and Mixed Noncompetitive?

Noncompetitive has equal affinity for E and ES, while Mixed has preference

What happens to enzyme’s affinity for substrate in Pure Noncompetitive Inhibition?

stays the same, affects catalysis not binding

What happens to Vmax in Pure Noncompetitive Inhibtion?

decreases

Why does Vmax change in Pure Noncompetitive Inhibition?

inhibitor removes active enzyme molecules from catalysis

What is Vmax apparent in Pure Noncompetitive Inhibition?

Vmax/a

What happens to the y-intercept in Pure Noncompetitive Inhibition?

becomes larger

What happens to the slope in Pure Noncompetitive Inhibition?

increases

What does a’ tell you?

how much inhibitor is bound to ES

What does it mean if alpha increases?

inhibitor binds more, substrate has harder time binding

What happens to Km if alpha increases?

increases

What happens to Vmax if alpha’ increases?

decreases

What happens to alpha and Ki if [Inhibitor Concentration] increases

alpha increases, Ki stays the same

What does alpha depend on?

both the inhibitor's binding strength and how much inhibitor is present

What happens to the x-intercept in Pure Noncompetitive Inhibition?

stays the same

What do you do if you see Pure Noncompetitive Inhibition?

test if it is a irreversible inhibitor thru dialysis

What two factors can you use to find alpha for Pure Noncompetitive Inhibition?

y-intercept and slope

How do you calculate Ki for Pure Noncompetitive Inhibition?

[I] / (m_i / m₀ - 1) OR [I] / (b_i / b₀ - 1)

What does a Mixed Noncompetitive Inhibitor bind do?

free enzyme OR enzyme-substrate complex (unequal affinity)

What does Ki’ tell you?

how tightly the inhibitor binds ES

Does Ki = Ki’ in Pure Noncompetitive Inhibition?

Yes, equal affinity for both E and ES

Does Ki = Ki’ in Mixed Noncompetitive Inhibition?

No, prefers E or ES

How many inhibition factors does are in Mixed Noncompetitive Inhibition?

Two (Ki, Ki’, a, a’)

What happens to Vmax in Mixed Noncompetitive Inhibition?

decreases

What does Km depend on in Mixed Noncompetitive Inhibition?

which the inhibitor prefers

What does inhibitor prefer if Ki < Ki’ ?

free enzyme

What does inhibitor prefer if Ki > Ki’ ?

enzyme substrate complex

What happens to Km if Ki < Ki’?

increases

What causes Km to change when Ki < Ki’?

inhibitor removes E from solution, substrate can’t find enzyme, enzyme appears to have lower affinity (higher Km)

What happens to Km if Ki > Ki’?

decreases

What causes Km to change when Ki > Ki’?

inhibitor binds ES tightly and stabilizes it, enzyme appears to bind substrate more

What happens to the slope in Mixed Noncompetitive Inhibition?

increases

What is the slope in Mixed Noncompetitive Inhibition?

aKm/Vmax

What happens to the y-intercept in Mixed Noncompetitive Inhibition?

increases (Vmax decreases)

Which ratio do you calculate alpha from for free enzyme in Mixed Noncompetitive Inhibition?

slope (m_i / m₀ )

Which ratio do you calculate alpha from for enzyme-substrate in Mixed Noncompetitive Inhibition?

y-intercept (b_i / b₀ )

How do you calculate a’?

y-int_i / y-int₀

Where do Mixed Inhibition lines intersect?

left of y-axis

Where do Mixed Inhibition lines intersect if prefer Enzyme?

above x-axis

Where do Mixed Inhibition lines intersect if prefer Enzyme-Substrate?

below x-axis

What does Uncompetitive Inhibitor bind to?

ES complex ONLY

What happens to Km in Uncompetitive Inhibition?

Km decreases

Why does Km change in Uncompetitive Inhibition?

inhibitor stabilizes ES complex, appears higher affinity

What happens to Vmax in Uncompetitive Inhibition?

Vmax decreases

Why does Vmax change in Uncompetitive Inhibition?

The inhibited complex ESI cannot make product

Can increasing substrate overcome Uncompetitive Inhibition?

No, will just create more ES for Inhibitor to bind

What is alpha equal to in Uncompetitive Inhibition?

a = 1, a’ > 1

What happens to the y-intercept in Uncompetitive Inhibition?

increases

What happens to the x-intercept in Uncompetitive Inhibition?

moves away from zero

What happens to the slope in Uncompetitive Inhibition?

stays the same, both Km and Vmax decrease by same factor

How do you calculate a’ in Uncompetitive Inhibition?

b_i / b₀

How do you calculate Ki’ in Uncompetitive Inhibition?

[I] / a’ - 1

What type of lines are shown in Uncompetitive Inhibition?

parallel lines

What is Vmax apparent in Uncompetitive Inhibition?

Vmax/a

What is Km apparent in Uncompetitive Inhibition?

Km/a

What does an Irreversible Inhibitor do?

permanently destroys enzyme activity

What happens to Vmax in Irreversible Inhibition?

decreases

What happens to Km in Irreversible Inhibition?

stays the same

What happens to y-intercept in Irreversible Inhibition?

increases

What happens to x-intercept in Irreversible Inhibition?

stays the same

What happens to slope in Irreversible Inhibition?

increases