This preview shows page 1. Sign up to view the full content.
Unformatted text preview: CEE 266
ENVIRONMENTAL BIOTECHNOLOGY
Lecture 3
(Enzyme Kinetics: MichaelisMenten equations,
LineweaverBurk plots, inhibition) Enzymes
Biological catalysts – increase the rate of a thermodynamicallyfavorable reaction
Literally “in yeast”
Proteins or RNA
Substrate specificity
Regenerated at the
end of reaction Six Classes of Enzymes Enzymes Need Support
Enzyme activity requires the native 3D
structure, and often the presence of a
cofactor such as a metal ion or
prosthetic group.
Apoenzymes lack the prosthetic group;
holoenzymes contain it and are fully
functional. Activation Energy and Catalysis Figure 5.6 Catalysis and Enzymes
Two mechanisms of catalysis EnzymeSubstrate Binding MichaelisMenten Kinetics
Rate of reaction depends on
Enzyme characteristics
Substrate Concentration
Affinity of enzyme for substrate
Formation of an enzymesubstrate complex
The ES complex is in rapid equilibrium with free enzyme
Breakdown of ES to form products is assumed to be slower than
formation of ES
breakdown of ES to reform E and S MichaelisMenten Kinetics Steady State v = k2[ES], if this is the ratelimiting step*
[Enzyme]total = [E]t = [E] + [ES]
1. Assume equilibrium, if k1 >> k2:
KS = k1/k1 = [E][S]/[ES]
or
2. Assume steady state:
d[ES]/dt = 0
[*Note: v is always measured as an initial rate!] MichaelisMenten Kinetics MichaelisMenten Kinetics Understanding Km
The ”affinity constant"
Km is a specific substrate concentration
Km is a constant derived from rate constants
Km is, under true MichaelisMenten conditions, an estimate of the
dissociation constant of E from S
Small Km means tight binding; high Km means weak binding Understanding Vmax
The theoretical maximal velocity
Vmax is a rate constant
Vmax is the theoretical maximal rate of the reaction  but it is NEVER
achieved in reality
To reach Vmax would require that ALL enzyme molecules are tightly
bound with substrate
Vmax is asymptotically approached as substrate is increased Dual Order Kinetics
Combination of 0order and 1storder kinetics
When S is low, the equation for rate is 1st order in S
When S is high, the equation for rate is 0order in S
The MichaelisMenten equation describes a rectangular hyperbolic
dependence of v on S! Double Reciprocal Plot Turnover Number Examples of Turnover Numbers Enzyme Affinity for Substrates Catalytic Efficiency, kcat/Km => “Perfect enzyme”
Diffusioncontrolled
limit: 108109 M1s1 An estimate of "how perfect" the enzyme is
kcat/Km is an apparent secondorder rate constant
It measures how the enzyme performs when S is low
The upper limit for kcat/Km is the diffusion limit  the rate at which E
and S diffuse together Types of Enzyme Inhibition Enzyme Inhibition Enzyme Inhibition Enzyme Inhibition Enzyme Inhibition Enzyme Inhibition Enzyme Inhibition Summary Example 1
MichaelisMenten kinetics can be used to describe the dependence of
biochemical reaction rate,V, on the concentration of substrate S.
a) For Vmax = 22 day1 and Km = 106 M, what is the growth rate at a
substrate concentration of 107 M. Example 1b b) If S were 10 times larger, how would this change your answer
quantitatively? c) If S were 1000 times larger, how would this change your answer
quantitatively? Please explain why, mechanistically, this would be
the case. Example 1c
If S were 1000 times larger, how would this change your answer
quantitatively? Please explain why, mechanistically, this would be
the case. Answers: V = 2 day1, @10*S, V = Vmax/2 = 11 day1, at
1000*S, V ~ Vmax = 22 day1. ...
View
Full
Document
This note was uploaded on 02/02/2012 for the course CEE 266 taught by Professor Shailymahendra during the Fall '11 term at UCLA.
 Fall '11
 ShailyMahendra
 Enzyme, Kinetics

Click to edit the document details