Enzyme kinetics practical

Question 1

What is Km?

Answer 1

A measure of enzyme’s affinity for a substrate, higher Km less affinity, slower reaction. Michaelis constant: the substrate concentration at which V (rate of reaction) is equal 1/2Vmax

Question 2

Variables affecting rate (initial rate)

Answer 2

Enzyme concentration [E] and also substrate concentration [S] but only until saturation reached and rate levels off at Vmax

Question 3

Michaelis menten equation

Answer 3

Gives the rate of reaction at a given substrate concentration.
v=Vmax[S]/([S]+Km)
Obviously only useful if you already know Vmax and Km

Question 4

Purpose of a lineweaver-burke plot

Answer 4

forms y=mx+c linear plot therefore allows you to graphically determine Vmax and Km.

Question 5

Lineweaver burke plot

Answer 5

Plot of 1/v against 1/[S]:
1/v = 1/Vmax + (Km/Vmax)(1/[S])
y = c + m x
==>c (intercept) = 1/Vmax
m (gradient) = Km/Vmax

Question 6

X-axis intercept of lineweaver burke plot

Answer 6

-1/Km

Question 7

Effect of Competitive inhibitor on Lineweaver Burke plot

Answer 7

Vmax stays the same (so y intercept 1/Vmax same). Apparent Km increases so gradient increases, x intercept changes.

Question 8

Effect of non-competitive inhibition on Lineweaver Burke plot

Answer 8

Reduces apparent Vmax so increases y intercept (1/Vmax).
Also increases gradient (Km/Vmax)
x-intercept stays the same as Km unnaffected

Question 9

Turnover number of an enzyme

Answer 9

maximum number of molecules of substrate that an enzyme can convert into product, per active site per second.