The role of electrostatic interactions in calmodulin-peptide complex formation.
The complex between calmodulin and the calmodulin-binding portion of smMLCKp has been studied. Electrostatic interactions have been anticipated to be important in this system where a strongly negative protein binds a peptide with high positive charge. Electrostatic interactions were probed by varying the pH in the range from 4 to 11 and by charge deletions in CaM and smMLCKp. The change in net charge of CaM from approximately -5 at pH 4.5 to -15 at pH 7.5 leaves the binding constant virtually unchanged. The affinity was also unaffected by mutations in CaM and charge substitutions in the peptide. The insensitivity of the binding constant to pH may seem surprising, but it is a consequence of the high charge on both protein and peptide. At low pH it is further attenuated by a charge regulation mechanism. That is, the protein releases a number of protons when binding the positively charged peptide. We speculate that the role of electrostatic interactions is to discriminate against unbound proteins rather than to increase the affinity for any particular target protein.
André
Ingemar
I
Department of Biophysical Chemistry, Lund University, Chemical Center, SE-22100 Lund, Sweden. ingemar.andre@bpc.lu.se
Kesvatera
Tõnu
T
Jönsson
Bo
B
Akerfeldt
Karin S
KS
Linse
Sara
S
D000595:Amino Acid Sequence; D000818:Animals; D002118:Calcium / Q000378:metabolism; D002147:Calmodulin / Q000737:chemistry*; D004926:Escherichia coli / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007477:Ions; D008024:Ligands; D008958:Models, Molecular; D015233:Models, Statistical; D008969:Molecular Sequence Data; D009010:Monte Carlo Method; D009154:Mutation; D010455:Peptides / Q000737:chemistry*; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011506:Proteins / Q000737:chemistry; D013050:Spectrometry, Fluorescence; D055672:Static Electricity; D013696:Temperature; D013816:Thermodynamics
D002147:Calmodulin; D007477:Ions; D008024:Ligands; D010455:Peptides; D011506:Proteins; D002118:Calcium