Catalytic efficiency has 2 components: kcat and Km.

kcat is the rate of catalytic turnover. Transition state theory proposes that kcat/kuncat is proportional to the affinity of the enzyme for the catalytic transition state, not the substrate.

Km is not the same as Kd except in the special case when the substrate is in rapid equilibrium with the enzyme relative to the rate of catalysis. When that is true, then the denominator of the catalytic efficiency is Kd, which decreases as the binding energy increases. Therefore, in that case catalytic efficiency increases with binding energy.

I think it would be possible to select mutants for enhanced binding affinity. However, you should regard information on binding energy from in silico studies as a hypothesis, which must be tested by experiment. In other words, make the mutants and measure the substrate binding affinity. (For a single-substrate enzyme, this may not be possible and you will have to measure the Km instead.)

In principle, the binding energy could be translated into the dissociation constant Kd. In practice, binding energies predicted by in silico techniques are probably not accurate enough. Moreover, Kd is often not equivalent to Km. Binding energy can not be used to predict kcat or kcat/Km, as far as I know.

Dear Adam,

Thank you somuch for your kind reply , As I understood you meant to say Its not possible to estimate catalytic efficency based on binding energy but I heard from some of colleques that catalytic efficiency is direct proportional to binding energy so is that correct?

 Also could you tell me , Suppose If my objective is to suggest a particular mutant for directed evolution based on binding energy  alone (highest) instead of catalytic efficiency. will it be possible?

Catalytic efficiency has 2 components: kcat and Km.

kcat is the rate of catalytic turnover. Transition state theory proposes that kcat/kuncat is proportional to the affinity of the enzyme for the catalytic transition state, not the substrate.

Km is not the same as Kd except in the special case when the substrate is in rapid equilibrium with the enzyme relative to the rate of catalysis. When that is true, then the denominator of the catalytic efficiency is Kd, which decreases as the binding energy increases. Therefore, in that case catalytic efficiency increases with binding energy.

I think it would be possible to select mutants for enhanced binding affinity. However, you should regard information on binding energy from in silico studies as a hypothesis, which must be tested by experiment. In other words, make the mutants and measure the substrate binding affinity. (For a single-substrate enzyme, this may not be possible and you will have to measure the Km instead.)

Dear Adam,

Thank you somuch for your kind reply.

I hope If I understood correct then you tried to say that predicting catalytic efficiency (kcat or Km) from binding energy is not possible but suggesting mutants for directed evolution based on binding energy is possible. This is the answer i expected but If possible could you suggest some articles regarding the same which would be really helpful me to make hypothesis

Thanking you in advance

It's not an area in which I can claim expertise. I found a very recent paper in PubMed that might provide some ideas and references. You should ask the corresponding author for a full-text copy.

http://www.ncbi.nlm.nih.gov/pubmed/25393978

Catalytic efficiency should be related to the binding energy of the transition state. Increasing binding energy for the ground state of the substrate will actually DECREASE catalytic rate.

Dear Adam,

Thank you somuch for your kind reply and I will let you know about the paper soon

Dear Roger,

Thanks for your kind reply but could you explain the same with detailed.

suppose, now I docked one enzyme with different mutant versions and I got binding energy  like below:

MutantPose;AFFINITY (kcal/mol);DistFromRmsd l.b.;BestModeRmsd u.b.

N4A.B99990001;-10.4;0.000;0.000

I2A.B99990001;-10.2;0.000;0.000

N4A.B99990001;-10.1;2.065;10.231

S1A.B99990001;-10.0;0.000;0.000

N3A.B99990001;-9.9;0.000;0.000

N4A.B99990001;-9.9;1.981;9.714

N3A.B99990001;-9.8;3.547;7.702

N4A.B99990001;-9.8;2.210;9.644

N3A.B99990001;-9.6;3.376;5.521

N4A.B99990001;-9.5;2.104;4.493

I2A.B99990001;-9.5;2.068;10.262

N3A.B99990001;-9.4;2.952;9.828

My question is how do you express highest binding energy in terms of catalytic efficiency ? whether it is highest/lowest catalytic efficiency ? I just want to know how we can relate this binding energy and catalytic efficiency in terms of docking result analysis.

I hope you will understand my doubt and thanking you in advance

As Roger said, kcat/Km measures the affinity to the transition state, not the ground state, so docking the substrate will not provide any insight to catalytic efficiency.  Often a very reactive substrate will have a high, not low, Km value, since it will react faster than it is released from the enzyme unreacted.  If you have a transition state analogue structure for the enzyme, docking that might give a better result, since it is known that transition state analogue affinity correlates better with kcat/Km than Km.

Rob

Robert is 100% correct.

To amplify a bit more, the free energy landscape from substrate to product can be "improved" in two ways to increase catalysis: you can lower the transition state free energy; you can raise the free energy of the bound substrate; or you can do both. The catalytic rate enhancement will depend on the decrease in free energy of activation between the bound substrate and the transition state. Measuring binding affinities of the substrate only gets you one part of the equation, and does not inform you of the other. In general (apply caveats mentioned above) tighter binding of the ground state (substrate) does not aid catalysis. (Many fast enzymes have rather high substrate Km values.) Tighter binding of the transition state--measurement of the binding of transition state analogs would be a reasonable proxy for this--is very likely to lead to more efficient catalysis, providing that the binding energy of the ground state substrate is also not tighter as well. (Then it's a wash.)

Thank you somuch all

 Quite apart from considerations of transition state etc., one very simple point that often gets overlooked is that every substrate that has to be bound has a corresponding product that has to be released and of course shares many/most of the structural features of the precursor substrate. If tight binding of the substrate also means tight binding of the product then product release could be rate limiting.

dear Roger,

thanks for the information provided so far but am still not clear. how will you measure the transition binding energy? and how will you relate it to the catalytic efficiency?