Enzyme practical Flashcards
What is the Michaelis Menten equation?
What are the units for [S] and v on the MM graph?
[S] = substrate –> mM
v = velocity –> uM/min
What is Kcat?
The turnover number.
It relates Vmax (he maximum velocity) to [Et], (the total active site concentration)
What is the equation for caculating the catalytic efficiency of an enzyme?
kcat /Km
the higher the value, the more efficient the enxyme is.
Describe the relationship between substrate and velocity in this graph.
[1] At very low [S]: Vo is proportional to [S] i.e.doubling [S] → double Vo.
[2] In mid-range of [S], Vo is increasing less as [S] increases.
Km = [S] when Vo = 1/2 Vmax.
[3] At very high [S], Vo is independent of [S]: Vo = Vmax.
What does ‘turnover number’ mean?
Number of substrate molecules converted into product by one molecule of enzyme active site per unit time.
Represented by k2.
Turnover number is Kcat.
also, V=K2[ES]
What is the equation for the Lineweaver Burk plot?
How can you calculate Km from this equation?
Km: an “aggregate” constant
(sum of rate constants for breakdown of ES divided by rate constant for formation of ES):
What is the equation for calculating Kcat?
Units are inverse time, e.g. s–1.
Fill in the axis for units and what is represented by the blue boxes.
How does a competitive inhibitor work?
How does it affect Vmax and Km values?
Competes with th SUBSTRATE for the enzyme active site.
Instead of E + S –> ES, you get E + I –> EI.
In the MM graph, Km increases, Vmax no change.
In the LB graph, 1/Km increases, 1/Vmax no change.
How does a non competitive inhibitor work?
How does it affect Vmax and Km values?
Competes with the ES complex, prevents the product being released.
Instead of ES –> E + P, you get
ES + I –> ESI.
In MM graph, Km no change, Vmax decreses.
In LB graph, 1/Km no change, 1/Vmax increases.
What is the equation for calculating Ki?
[I] = conc of inhibitor.
Km(app) = Km with the inhibitor.
What equation would you use if you wanted to estimate the value of Km on a graph with
[S] on the x-axis and
velocity on the y-axis?
Km = 1/2 Vmax.
How does a Non-competetive enzyme inhibitor work?
A non-competitive inhibitors binds to a site other than the active site.
The enzyme can have both the substrate and inhibitor bound at the same time.
ES + I –> ESI
The the enzyme-substrate complex can still form and there is no reduced affinity for the substrate.
Km stays the same.
The enzyme-substrate complex cannot make any product, so the Vmax is lowered.
