Hi Francis I guess it depends on how much residual high-affinity binder you have in the mixture and what the difference in affinity is between Y and deriv-Y. Another issue is of course whether Y and derY compete for the same binding site and have the same stoichiometry. A well designed displacement ITC experiment and comparisons thereof with ITC data for your high-affinity binder should lead to some good answers. Knowing the ratio of Y vs deriv-Y in your starting compound solution will be an advantage.
A very useful reference in thinking about and carrying out displacement ITC in our group has been the one by Velazquez-Campoy and Freire. This article was specifically written to address the application of displacement titrations in ITC. We have applied this approach to address several types of questions concerning interactions in the uM-pM range. Velazquez-Campoy A, Freire E. Isothermal titration calorimetry to determine association constants for high-affinity ligands Nat Protoc. 2006;1(1):186-91. Best regards Savvas ---- Savvas Savvides Unit for Structural Biology @ L-ProBE Ghent University K.L. Ledeganckstraat 35, 9000 Ghent, Belgium Ph. +32 (0)472 928 519 http://www.LProBE.ugent.be/xray.html On 24 Aug 2010, at 17:11, Francis E Reyes wrote: > Hi All > > I'm curious the effect of small impurities in commercially synthesized > compounds on ITC and its analysis. Say if compound Y is the high affinity > binder, but you make a derivative that differs from a single functional group > from Y (you used Y to make this new compound) and you never are able to > completely get rid of Y. How does this affect the analysis of determining the > derivative's affinity by ITC? > > References or personal experience is appreciated! > > F > > --------------------------------------------- > Francis E. Reyes M.Sc. > 215 UCB > University of Colorado at Boulder > > gpg --keyserver pgp.mit.edu --recv-keys 67BA8D5D > > 8AE2 F2F4 90F7 9640 28BC 686F 78FD 6669 67BA 8D5D
