Dear Sajid If the binding site of your ligand is remote from the dimerization interface, it should normally not be a problem. You will bind two ligands for a dimer and one ligand for a monomer and you should be able to fit the isotherm with one unique site even in presence of a mix of monomers and dimers.
If the binding site is close to the dimerization interface or partially dissociate the dimer, this may give more complicate signals difficult to analyzed You can also try to evaluate with ITC or other biophysical technic, the Kd of your dimer. With ITC, the Kd of your dimer may be evaluated by injecting a concentrated solution of your protein via the serynge against a cell containing your dialysis buffer. You may observe an heat exchange due to dimer dissociation (see for example Li, J, Weis RM, 1996, with CheA) Knowing the Kd may help to design the ITC experiment. For example, you can use a concentration of protein 10 fold higher (if possible) than the Kd to have mainly dimers Cheers JB JB Charbonnier Laboratory of Structural Biology and Radiobiology iBiTec-S (Institute of Biology and Technologies of Saclay), CEA , FRANCE [email protected] -----Message d'origine----- De : CCP4 bulletin board [mailto:[email protected]] De la part de sajid akthar Envoyé : vendredi 18 juillet 2014 11:24 À : [email protected] Objet : [ccp4bb] ITC with heterogeneous protein 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
