| 研究実績の概要 |
Cholix toxin (ChxA) is a recently identified cytotoxin produced by pathogenic Vibrio cholerae non-O1/non-O139. The cytotoxic pathway and mouse lethality caused by liver injury has been gradually understood, however, the cell surface receptors are not yet known, which are necessary molecules for ChxA binding to target cells. The objectives for the fiscal year 2023-24 were focused on attempting to purify the recombinant candidate receptor protein (cRP) by solubilizing recombinatn Spodoptera frugiperda (Sf9) cells carrying the gene for the histidine-tagged cRP, using 8M urea and purifying the cRP. Additionally, the goal was to achieve direct binding between ChxA and cRP.
Unsuccessful purification attempts: Despite extensive efforts, the purification of cRP could not be achieved using the proposed methods. Solubilization of whole cell proteins with 8M urea followed by nickel affinity chromatography failed to yield purified cRP. The inability to purify cRP suggests challenges related to its large size or other unknown factors.
Failure to establish direct binding: Due to the unsuccessful purification of cRP, direct binding studies between ChxA and cRP could not be conducted. The lack of binding observed in previous attempts also indicates a barrier to establishing this interaction.
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