Abstract
The voltage-gated proton channel Hv1 functions as a dimer, in which the intracellular C-terminal domain of the protein is responsible for the dimeric architecture and regulates proton permeability. Although it is well known that divalent metal ions have effect on the proton channel activity, the interaction of divalent metal ions with the channel in detail is not well elucidated. Herein, we investigated the interaction of divalent metal ions with the C-terminal domain of human Hv1 by CD spectra and fluorescence spectroscopy. The divalent metal ions binding induced an obvious conformational change at pH 7 and a pH-sensitive reduction of thermostability in the C-terminal domain. The interactions were further estimated by fluorescence spectroscopy experiments. There are at least two binding sites for divalent metal ions binding to the C-terminal domain of Hv1, either of which is close to His244 or His266 residue. The binding of Zn2+ to the two sites both enhanced the fluorescence of the protein at pH 7, whereas the binding of other divalent metal ions to the two sites all resulted fluorescence quenching. The orders of the strength of divalent metal ions binding to the two sites from strong to weak are both Co2+, Ca2+, Ni2+, Mg2+, and Mn2+. The strength of Ca2+, Co2+, Mg2+, Mn2+ and Ni2+ binding to the site close to His244 is stronger than that of these divalent metal ions binding to the site close to His266.
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Acknowledgments
This work was supported by National Natural Science Foundation of China (No. 30970579 and 31271464), and the Ph.D. Programs Foundation of Ministry of Education of China (No. 20110031110004 and 20120031110028), and the Basic Science and Advance Technology Research Program of Tianjin (No. 14JCYBJC23400).
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Zhao, Q., Zhang, Y. & Li, S.J. Interaction of divalent metal ions with the carboxyl-terminal domain of human voltage-gated proton channel Hv1. Biometals 27, 793–802 (2014). https://doi.org/10.1007/s10534-014-9751-6
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DOI: https://doi.org/10.1007/s10534-014-9751-6