chem axon is used for the predict ion of dissociation constant

I think, in few docking programs you can make few bonds rigid to keep the desired steriochemistry and carry out docking with flexible or rigid active site. By this you could get dissociation constant, binding free energy calculation seperatly for both the isomers. As there are two binding sites you have to again find the best one which gives the lowest binding free enegy.

I think, in few docking programs you can make few bonds rigid to keep the desired steriochemistry and carry out docking with flexible or rigid active site. By this you could get dissociation constant, binding free energy calculation seperatly for both the isomers. As there are two binding sites you have to again find the best one which gives the lowest binding free enegy.

Hi Lakshmi, 

I think docking is a good tool provided you use it wisely! It might give you the right answers, or it could lead you down a rabbit hole. As Rajesh has pointed out, you can certainly model isomers, and compare binding. But an experimental result will be of far greater value, provided it is possible to carry out.

I am curious about why you have the 80-20 mixture of isomers. Is it due to an unavoidable equilibrium, or a result of a particular synthesis method? Can the isomers exist separately?

@ Nitesh Sule I am trying to repurpose the drug, an antibiotic, which is produced by the microbe as a mixture of these compounds which differ only by a methyl group. The drug is already proven safe for use in humans and it shows potent cytotoxic activity,which is a novel finding. I am trying to elucidate the mechanism by which it kills the cells. 

It is binding to the protein of interest but I am unable to calculate its Kd because even the overall fluorescence is enhanced at all concentrations of the drug, the trend is not linear. Could it be due to the differential binding of the 2 ligands? 

Good line of work @ Laxmi..If the Kd /Ki values of the off target effects are experimentally available along with the docking studies, MD studies could be more accurate and realistic. Further, is there any similarity between the active sites of proteins you are targeting?  This could explain or give you some hints of binding modes of two isomers.

Best of luck..

That is certainly interesting.

Allow me to step back a little, and ask you - are these constitutional isomers (2-methyl-drug vs 3-methyl-drug) or stereoisomers (R-3-methyl-drug vs S-3-methyl-drug or something like that)? In either case, they can bind differently to the receptor, but I'm curious if they are produced by the microbe via independent pathways, or if its a single product that is enzymatically isomerized. Depending on the answer, and the project goals, you might consider a slightly different approach to your experiments.

Coming back to the immediate problem - the difficulty in determining the Kd. I am afraid I did not quite follow your description of the situation (the overall fluorescence is enhanced at all concentrations of the drug, the trend is not linear). Could you perhaps post a figure or drawing to explain what you mean? If you have two isomeric ligands and suspect two binding sites, then the situation could be quite complex.