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Title: Identification and In-vivo Characterization of a Novel OhrR Transcriptional Regulator in Burkholderia xenovorans LB400

Transcriptional regulators (TRs) are an important and versatile group of proteins, yet very little progress has been achieved towards the discovery and annotation of their biological functions. We have characterized a previously unknown organic hydroperoxide resistance regulator from Burkholderia xenovoransLB400, Bxe_B2842, which is homologous to E. coli’s OhrR. Bxe_B2842 regulates the expression of an organic hydroperoxide resistance protein (OsmC). We utilized frontal affinity chromatography coupled with mass spectrometry (FAC-MS) and electrophoretic mobility gel shift assays (EMSA) to identify and characterize the possible effectors of the regulation by Bxe_B2842. Without an effector, Bxe_B2842 binds a DNA operator sequence (DOS) upstream of osmC. FAC-MS results suggest that 2-aminophenol binds to the protein and is potentially an effector molecule. EMSA analysis shows that 2-aminophenol also attenuates the Bxe_B2842’s affinity for its DOS. EMSA analysis also shows that organic peroxides attenuate Bxe_B2842/DOS affinity, suggesting that binding of the TR to its DOS is regulated by the two-cysteine mechanism, common to TRs in this family. Bxe_B2842 is the first OhrR TR to have both oxidative and effector-binding mechanisms of regulation. Our paper reveals further mechanistic diversity TR mediated gene regulation and provides insights into methods for function discovery of TRs.
Authors:
 [1] ;  [1] ;  [1] ;  [1] ;  [2] ;  [1] ;  [1] ;  [1]
  1. Los Alamos National Lab. (LANL), Los Alamos, NM (United States)
  2. Los Alamos National Lab. (LANL), Los Alamos, NM (United States)ry
Publication Date:
OSTI Identifier:
1224042
Report Number(s):
LA-UR--13-21680
Journal ID: ISSN 1925-430X
Grant/Contract Number:
AC52-06NA25396
Type:
Accepted Manuscript
Journal Name:
Journal of Molecular Biology Research
Additional Journal Information:
Journal Volume: 3; Journal Issue: 1; Journal ID: ISSN 1925-430X
Publisher:
Canadian Center of Science and Education
Research Org:
Los Alamos National Lab. (LANL), Los Alamos, NM (United States)
Sponsoring Org:
USDOE
Country of Publication:
United States
Language:
English
Subject:
59 BASIC BIOLOGICAL SCIENCES transcriptional regulator (TR), transcriptional regulator effector (TRE); fontal affinity chromatography coupled to mass spectrometry (FAC-MS), Burkholderia xenovorans LB400; organic hydroperoxide resistance regulator (OhrR); derepression; organic peroxides; Bxe_B2842 and 2-aminophenol