Mass Spectrometric Determination of Zn 2+ Binding/Dissociation Constant for Zinc Finger Peptides

Abstract

Abstract : In the present study, we proposed a simple ESI-MS model for determining Zn 2+ binding (or dissociation) constants forzinc finger peptides (ZFPs) with a unique ββα fold consensus. The ionization efficiency (response) factors for this model, i.e., α andβ, could be determined for ZiCo ZFP with a known Zn 2+ binding constant. We could determine the binding constants for other ZFPsassuming those with a ββα consensus conformation have the same α/β response ratio. In general, the ZPF dissociation constantsexhibited K d values of 10 -7 ~10 -9 M, while K d values for a negative control non-specific Zn 2+ peptides were high, e.g., 5.5 ×10 -6 M and4.3×10 -4 M for BBA1 and melittin, respectively.Key words : zinc finger, binding constant, electrospray-mass spectrometry, zinc ion Introduction Since its discovery in transcription factor IIIA (TFIIIA)from Xenopuslaevis, zinc finger proteins (ZFPs) have beenextensively researched due to their implications foreukaryotic protein-nucleic acid interactions.