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I am investigating the strength of binding of a small peptide to a protein by isolating the bound version and subjecting it to collisions with gas molecules (CID mass spectrometry) to dissociate the complex. When I plot the collision energy against the proportion of bound and unbound proteins I get a curve that looks like a Kd curve. However, as the interaction isn't in equilibria (i.e. it is in the gas phase) calculating the Kd value wouldn't be correct.

So is there another value I could calculate to describe the strength of the interaction?

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up vote 3 down vote accepted

Bobthejoe was pointing in the right direction kcat and Km - Michaelis Menton parameters are measured to look at the relative rates of an enzyme in non equilibrium state. Basically kcat is 'how fast does the enzyme work with little or no product in solution'. Km is 'how strongly does the enzyme bind the substrates that it uses.

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You should be looking at the interactions in terms of kinetics and not thermodynamics. k_on and k_off should do the trick.

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I don't think this is right because, k-on/k-off = Ka and Ka=1/Kd. So that would mean having K-on and K-off would allow you to calculate Kd, which is an equilibrium measurement... ah well thanks for trying! – Anake Mar 2 '12 at 12:02
Your basing your argument on psuedosteady state criteria and a constant concentration of your peptides and proteins. k_on and k_off are exclusively kinetic based terms that can be extended to thermodynamic terms when in equilibria which you're not in. – bobthejoe Mar 2 '12 at 23:08

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