title: Experimental and calculated shift in pK(a) upon binding of phosphotyrosine peptide to the SH2 domain of p56(lck).
abstract: The pH dependence of the affinity of a 11-mer phosphotyrosine (pY) peptide (EPQpYEEIPIYL-NH2) for the SH2 domain of the tyrosine kinase p56(lck) was investigated with surface plasmon resonance (SPR). From SPR competition experiments the affinity in solution was obtained. The pH dependence of the affinity in solution can be well described by a proton linkage model with a single pK(a) shift upon binding, from 6.1 to 4.7. This shift is ascribed to the transition from the -2 to the -1 ionisation state of the tyrosine phosphate group. Based on the X-ray structure for the complex with Lck SH2, a pK(a) value of 5.3 for the bound pY peptide was computed, modelling the solvated protein as a system of point charges in a continuum. With the phosphate in the -2 state the binding energy is 1.8 kcal/mol more favourable than for the -1 state, corresponding to a 20-fold higher affinity. A proper charge is relevant in the design of potential therapeutic Lck SH2 ligands with mimics for the metabolically unstable tyrosine phosphate group.
authors.affiliation: Department of Medicinal Chemistry, Utrecht Institute for Pharmaceutical Sciences, Faculty of Pharmacy, Utrecht University, PO Box 80082, 3508TB, Utrecht, The Netherlands. n.j.demol@pharm.uu.nl
authors.forename: Nico J
authors.initials: NJ
authors.lastname: de Mol
authors.forename: Malcolm B
authors.initials: MB
authors.lastname: Gillies
authors.forename: Marcel J E
authors.initials: MJ
authors.lastname: Fischer
mesh_terms: D001665:Binding Sites; D015195:Drug Design; D006801:Humans; D006863:Hydrogen-Ion Concentration; D008024:Ligands; D019860:Lymphocyte Specific Protein Tyrosine Kinase p56(lck) / Q000378:metabolism*; D008956:Models, Chemical; D009842:Oligopeptides / Q000737:chemistry / Q000378:metabolism*; D019000:Phosphotyrosine / Q000737:chemistry*; D011485:Protein Binding; D012996:Solutions; D020349:Surface Plasmon Resonance; D013816:Thermodynamics; D018909:src Homology Domains
chemical_list: D008024:Ligands; D009842:Oligopeptides; D012996:Solutions; D019000:Phosphotyrosine; D019860:Lymphocyte Specific Protein Tyrosine Kinase p56(lck)