Crystal Structure of the Regulatory Domain of AphB from Vibrio vulnificus, a Virulence Gene Regulator

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.

iDEP: an integrated web application for differential expression and pathway analysis of RNA-Seq data

“…Applying PGSEA on miRNA target genes highlights miRNA-30a (Additional file 3: Figure S20), whose target genes are specifically activated by IR in wild-type B cells. miRNA-30a was shown to be involved in response to IR [77] and mutually regulate p53 [78]. Thus, the complex p53 signaling pathways are unveiled with remarkable accuracy.

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The Global Acetylome of the Human Pathogen Vibrio cholerae V52 Reveals Lysine Acetylation of Major Transcriptional Regulators

“…Both residues are found far from the DNA binding site, but while K271 is located on the back relative to the dimerization interface, the K103 was in a position to potentially affect dimerization (Figure 7A ). In the Vibrio vulnificus AphB homolog, K103 was shown to interact with an as yet unknown ligand (Park et al, 2017 ). Acetylation of vibriobactin transporter VctP (KNH52012) K110 was identified in both growth conditions.…”

STRUCTURAL STUDIES OF TRANSCRIPTIONAL REGULATION BY LysR-TYPE TRANSCRIPTIONAL REGULATORS IN BACTERIA

“…The RD of LTTRs has an inducer binding cavity (IBC) and is presumed to play a critical role in the conformational change of the LTTR tetramer upon inducer binding (Choi et al, 2001;Maddocks and Oyston, 2008;Quade et al, 2011;Park et al, 2017). The RD from CysB was the first crystal structure solved of a RD (Tyrell et al, 1997).…”

“…RD-II contains a five-stranded -sheet that is strongly twisted and four -helices ( Fig. 4) (Tyrell et al, 1997;Muraoka et al, 2003;Monferrer et al, 2010;Quade et al, 2011;Park et al, 2017). Structural studies of BenM, OxyR, PcaQ, RovM, AphB and DntR have led us to hypothesize that inducer binding (or environmental change) to the RD of LTTR causes a conformational change in the RD that is propagated throughout the tetrameric LTTR and changes the bend angle of the promoter DNA (Kovacikova and Skorupski, 2001;Bundy et al, 2002;Smirnova et al, 2004;Quade et al, 2011;Wei et al, 2012;Jo et al, 2015) However, while crystal structures of OxyR (Choi et al, 2001;Jo et al, 2015), BenM (Ezezika et al, 2007a) and DntR (Devesse et al, 2011) have revealed conformational changes of the RD upon inducer binding, conformational changes of tetrameric full-length LTTR upon inducer binding have not been observed in the crystal.…”

“…In the closed form, there are interactions between two helices from two RD-II subdomains (two V helices from two distinct RD-II) that are related by a two-fold axis. Upon inducer binding, local conformational changes in the RD (Ezezika et al, 2007a;Devesse et al, 2011;Park et al, 2017) seem to disrupt helix-helix interactions leading to a structural change to the open form ( Fig. 3(C)) (Choi et al, 2001;Monferrer et al, 2010;Devesse et al, 2011).…”