HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS		QUICK SEARCH:   [advanced] Author: Keyword(s): Year:  Vol:  Page:

J. Biol. Chem., Vol. 281, Issue 33, 23567-23578, August 18, 2006

This Article Full Text Full Text (PDF) Supplemental Data All Versions of this Article: 281/33/23567    most recent M603968200v1 Submit a Letter to Editor Alert me when this article is cited Alert me when eLetters are posted Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Request Permissions Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Lee, J.-W. Articles by Helmann, J. D. Search for Related Content PubMed PubMed Citation Articles by Lee, J.-W. Articles by Helmann, J. D. Social Bookmarking                      What's this?

Biochemical Characterization of the Structural Zn²⁺ Site in the Bacillus subtilis Peroxide Sensor PerR^*

From the Department of Microbiology, Cornell University, Ithaca, New York 14853-8101

In Bacillus subtilis most peroxide-inducible oxidative stress genes are regulated by a metal-dependent repressor, PerR. PerR is a dimeric, Zn²⁺-containing metalloprotein with a regulatory metal-binding site that binds Fe²⁺ (PerR:Zn,Fe) or Mn²⁺ (PerR: Zn,Mn). Reaction of PerR:Zn,Fe with low levels of hydrogen peroxide (H₂O₂) leads to oxidation of two His residues thereby leading to derepression. When bound to Mn²⁺, the resulting PerR:Zn,Mn is much less sensitive to oxidative inactivation. Here we demonstrate that the structural Zn²⁺ is coordinated in a highly stable, intrasubunit Cys₄:Zn²⁺ site. Oxidation of this Cys₄:Zn²⁺ site by H₂O₂ leads to the formation of intrasubunit disulfide bonds. The rate of oxidation is too slow to account for induction of the peroxide stress response by micromolar levels of H₂O₂ but could contribute to induction under severe oxidative stress conditions. In vivo studies demonstrated that inactivation of PerR:Zn,Mn required 10 mM H₂O₂, a level at least 1000 times greater than that needed for inactivation of PerR:Zn,Fe. Surprisingly even under these severe oxidation conditions there was little if any detectable oxidation of cysteine residues in vivo: derepression was correlated with oxidation of the regulatory site. Because oxidation at this site required bound Fe²⁺ in vitro, we suggest that treatment of cells with 10 mM H₂O₂ released sufficient Fe²⁺ into the cytosol to effect a transition of PerR from the PerR:Zn,Mn form to the peroxide-sensitive PerR: Zn,Fe form. This model is supported by metal ion affinity measurements demonstrating that PerR bound Fe²⁺ with higher affinity than Mn²⁺.

Received for publication, April 25, 2006 , and in revised form, June 6, 2006.

^* This work was supported by National Science Foundation Grant MCB-0235255. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

The on-line version of this article (available at http://www.jbc.org) contains supplemental Figs. S1 and S2.

¹ To whom correspondence should be addressed: Dept. of Microbiology, 370 Wing Hall, Cornell University, Ithaca, NY 14853. Tel.: 607-255-6570; Fax: 607-255-3904; E-mail: jdh9{at}cornell.edu.

CiteULike    Complore    Connotea    Del.icio.us    Digg    Reddit    Technorati    What's this?

This article has been cited by other articles:

				E. S. Anderson, J. T. Paulley, J. M. Gaines, M. W. Valderas, D. W. Martin, E. Menscher, T. D. Brown, C. S. Burns, and R. M. Roop II The Manganese Transporter MntH Is a Critical Virulence Determinant for Brucella abortus 2308 in Experimentally Infected Mice Infect. Immun., August 1, 2009; 77(8): 3466 - 3474. [Abstract] [Full Text] [PDF]

				L. I. Salzberg and J. D. Helmann An Antibiotic-Inducible Cell Wall-Associated Protein That Protects Bacillus subtilis from Autolysis J. Bacteriol., July 1, 2007; 189(13): 4671 - 4680. [Abstract] [Full Text] [PDF]

				J.-W. Lee, S. Soonsanga, and J. D. Helmann A complex thiolate switch regulates the Bacillus subtilis organic peroxide sensor OhrR PNAS, May 22, 2007; 104(21): 8743 - 8748. [Abstract] [Full Text] [PDF]

				M. Leelakriangsak, K. Kobayashi, and P. Zuber Dual Negative Control of spx Transcription Initiation from the P3 Promoter by Repressors PerR and YodB in Bacillus subtilis J. Bacteriol., March 1, 2007; 189(5): 1736 - 1744. [Abstract] [Full Text] [PDF]