it depend of what you expected as information : In this case your measure is resulting from ligand binding AND dimerization (except if all the protein is already dimerized). I am not sur to understand, do you know how many binding sites exists on one monomer ? You should be able to determine the stoechiometry ligand/protein, but it's not necessarily the same between monomer and dimer. Is the ligand "trigger" for dimerization ? Or the dimerization appear in all case upon the concentration is sufficient ?
________________________________________ De : CCP4 bulletin board [CCP4BB@JISCMAIL.AC.UK] de la part de Ramesh V [ramesh.c...@gmail.com] Envoyé : vendredi 18 juillet 2014 14:42 À : CCP4BB@JISCMAIL.AC.UK Objet : Re: [ccp4bb] ITC with heterogeneous protein I have a similar case, where in there are multiple binding sites on the protein for the ligand and ligand induces dimerization.So it is not helpful even if I separate the monomer and dimer. If I titrate the dimer with ligand, the stoichiometry will completely change? Any suggestions will be helpful. On Fri, Jul 18, 2014 at 12:46 PM, David Briggs <drdavidcbri...@gmail.com<mailto:drdavidcbri...@gmail.com>> wrote: Hi Sajid, *Assuming* you have one site per monomer (rather than, say, one site per dimer), and *assuming* each binding event is completely independent ( I.e no co-operativity), you might just get away with running the experiment with the heterogeneous material. However, you might not be able to confidently make these assumptions, so imho it would be preferable to separate the monomer and dimer by SEC prior to ITC. If this is not possible, then pay close attention to the fit when you run the heterogeneous experiment. Poor fit to a one site model may indicate that these assumptions are invalid. Can you obtain stoichiometry information from a different technique? This might be very helpful. Hth, Dave Dr David C Briggs PhD http://about.me/david_briggs On 18 Jul 2014 10:25, "sajid akthar" <b_sajid_...@yahoo.co.in<mailto:b_sajid_...@yahoo.co.in>> wrote: Dear All, This is an off-topic question. I have protein solution of heterogeneous (contains both monomer and dimer). I want to perform ITC with this protein. I doubt whether this heterogeneity will interfere the binding study. Any advice please. Thank you Sajid
