James- It's: lnKd = deltaH- RT(deltaS) As for Km ~ Kd, here be dragons...Km values cannot be treated as the Kd of a complex because dissociation of a catalytic complex has two potential fates, one that is denoted by a rate constant that is correlated to turnover (so, the forward reaction leading to product, generically called k2) and one that represents the degeneracy or breakdown of the enzyme/substrate complex (the reverse reaction or k-1). The Km will most likely be very similar to the Kd but it is only an apparent Kd (Kdapp). Since you have measured enzyme activities at multiple temperatures, you could graph an Arrhenius plot and determine the activation free energies for the wt and mutant enzymes by a statistical fit to the plotted data. This would provide a comparative thermodynamic analysis of the enzyme. If you have a good substrate analog (inhibitor), you could determine Ki with your normal activity assay and compare between wt and mutant. Of course, there's always calorimetry, too...
HTH- Brad On Wed, Nov 18, 2009 at 11:24 AM, james09 pruza <[email protected]>wrote: > Dear All, > Sorry for the non-ccp4 query. > I have solved a crystal structure of an enzyme and woring on its > biochemical aspect. We have a mutant of this enzyme and we are comparing > some thermodynamic parameters of this enzyme with mutant( lke delH and delS, > delG). we have done the expt at different temp. and know the km value at > these temp. Now the question is how to get the value of delH and delS. > The relation of these parameters is:- > -RTlnK = delH - TdelS > > Is the K in this relation is km or kd. If it can be treated as kd, so it > should be 1/km. > > All suggestions are welcome. > Thanks > James >
