Construction of a Ca²⁺-Gated Artificial Channel by Fusing Alamethicin with a Calmodulin-Derived Extramembrane Segment

Abstract

Using native chemical ligation, we constructed a Ca²⁺-gated fusion channel protein consisting of alamethicin and the C-terminal domain of calmodulin. At pH 5.4 and in the absence of Ca²⁺, this fusion protein yielded a burst-like channel current with no discrete channel conductance levels. However, Ca²⁺ significantly lengthened the specific channel open state and increased the mean channel current, while Mg²⁺ produced no significant changes in the channel current. On the basis of 8-anilinonaphthalene-1-sulfonic acid (ANS) fluorescent measurement, Ca²⁺-stimulated gating may be related to an increased surface hydrophobicity of the extramembrane segment of the fusion protein