Crystal Structure of the Regulatory Domain of AphB from Vibrio vulnificus, a Virulence Gene Regulator
Nohra Park1,3, Saemee Song1,3, Garam Choi1,2, Kyung Ku Jang1,2, Inseong Jo1, Sang Ho Choi1,2,*, and
Nam-Chul Ha1,*
1Department of Agricultural Biotechnology, Center for Food Safety and Toxicology, and Research Institute for Agriculture and
Life Sciences, Seoul National University, Seoul 08826, Korea, 2National Research Laboratory of Molecular Microbiology and Toxicology,
Seoul National University, Seoul 08826, Korea, 3These authors contributed equally to this work.
*Correspondence: choish@snu.ac.kr (SHC); hanc210@snu.ac.kr (NCH)
Received February 2, 2017; Revised April 5, 2017; Accepted April 7, 2017.; Published online April 20, 2017.
© Korean Society for Molecular and Cellular Biology. All rights reserved.

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ABSTRACT
The transcriptional activator AphB has been implicated in acid resistance and pathogenesis in the food borne pathogens Vibrio vulnificus and Vibrio cholerae. To date, the full-length AphB crystal structure of V. cholerae has been determined and characterized by a tetrameric assembly of AphB consisting of a DNA binding domain and a regulatory domain (RD). Although acidic pH and low oxygen tension might be involved in the activation of AphB, it remains unknown which ligand or stimulus activates AphB at the molecular level. In this study, we determine the crystal structure of the AphB RD from V. vulnificus under aerobic conditions without modification at the conserved cysteine residue of the RD, even in the presence of the oxidizing agent cumene hydroperoxide. A cysteine to serine amino acid residue mutant RD protein further confirmed that the cysteine residue is not involved in sensing oxidative stress in vitro. Interestingly, an unidentified small molecule was observed in the inter-subdomain cavity in the RD when the crystal was incubated with cumene hydroperoxide molecules, suggesting a new ligand-binding site. In addition, we confirmed the role of AphB in acid tolerance by observing an aphB-dependent increase in cadC transcript level when V. vulnificus was exposed to acidic pH. Our study contributes to the understanding of the AphB molecular mechanism in the process of recognizing the host environment.
Keywords: crystal structure, low pH, transcriptional regulator
AphB, vibrio vulnificus


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