Vibrio campbellii chitoporin: Thermostability study and implications for the development of therapeutic agents against Vibrio infections

Abstract

Vibrio campbellii (formerly Vibrio harveyi) is a bacterial pathogen that causes vibriosis, which devastates fisheries and aquaculture worldwide. V. campbellii expresses chitinolytic enzymes and chitin binding/transport proteins, which serve as excellent targets for antimicrobial agent development. We previously characterized VhChiP, a chitooligosaccharide-specific porin from the outer membrane of V. campbellii BAA-1116. This study employed far-UV circular dichroism and tryptophan fluorescence spectroscopy, together with single channel electrophysiology to demonstrate that the strong binding of chitoligosaccharides enhanced thermal stability of VhChiP. The alanine substitution of Trp136 at the center of the affinity sites caused a marked decrease in the binding affinity and decreased the thermal stability of VhChiP. Tryptophan fluorescence titrations over a range of temperatures showed greater free-energy changes on ligand binding (ΔG°binding) with increasing chain length of the chitooligosaccharides. Our findings suggest the possibility of designing stable channel-blockers, using sugar-based analogs that serve as antimicrobial agents, active against Vibrio infection.