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

J. Biol. Chem., Vol. 281, Issue 44, 33152-33162, November 3, 2006

This Article Full Text Full Text (PDF) Supplemental Data All Versions of this Article: 281/44/33152    most recent M606245200v1 Alert me when this article is cited 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 Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Login, F. H. Articles by Shevchik, V. E. Search for Related Content PubMed PubMed Citation Articles by Login, F. H. Articles by Shevchik, V. E. Social Bookmarking                      What's this?

The Single Transmembrane Segment Drives Self-assembly of OutC and the Formation of a Functional Type II Secretion System in Erwinia chrysanthemi^*

From the Unité de Microbiologie et Génétique, UMR 5122 CNRS, INSA de Lyon, Université Lyon 1, 69622 Villeurbanne, France

Many pathogenic Gram-negative bacteria secrete toxins and lytic enzymes via a multiprotein complex called the type II secretion system. This system, named Out in Erwinia chrysanthemi, consists of 14 proteins integrated or associated with the two bacterial membranes. OutC, a key player in this process, is probably implicated in the recognition of secreted proteins and signal transduction. OutC possesses a short cytoplasmic sequence, a single transmembrane segment (TMS), and a large periplasmic region carrying a putative PDZ domain. A hydrodynamic study revealed that OutC forms stable dimers of an elongated shape, whereas the PDZ domain adopts a globular shape. Bacterial two-hybrid, cross-linking, and pulldown assays revealed that the self-association of OutC is driven by the TMS, whereas the periplasmic region is dispensable for self-association. Site-directed mutagenesis of the TMS revealed that cooperative interactions between three polar residues located at the same helical face provide adequate stability for OutC self-assembly. An interhelical H-bonding mediated by Gln²⁹ appears to be the main driving force, and two Arg residues located at the TMS boundaries are essential for the stabilization of OutC oligomers. Stepwise mutagenesis of these residues gradually diminished OutC functionality and self-association ability. The triple OutC mutant R15V/Q29L/R36A became monomeric and nonfunctional. Self-association and functionality of the triple mutant were partially restored by the introduction of a polar residue at an alternative position in the interhelical interface. Thus, the OutC TMS is more than just a membrane anchor; it drives the protein self-association that is essential for formation of a functional secretion system.

Received for publication, June 29, 2006 , and in revised form, August 11, 2006.

^* This work was supported by grants from CNRS, INSA de Lyon, and the French Ministry of Research and Technology. 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 and Table S1.

¹ To whom correspondence should be addressed: UMG UMR 5122 CNRS-INSA-UCB, Bat. Lwoff, 10 rue Dubois, 69622 Villeurbanne France. Tel.: 33-472-445-827; Fax: 33-472-431-584; E-mail: vladimir.shevchik{at}insa-lyon.fr.

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

This article has been cited by other articles:

				C. Creze, S. Castang, E. Derivery, R. Haser, N. Hugouvieux-Cotte-Pattat, V. E. Shevchik, and P. Gouet The Crystal Structure of Pectate Lyase PelI from Soft Rot Pathogen Erwinia chrysanthemi in Complex with Its Substrate J. Biol. Chem., June 27, 2008; 283(26): 18260 - 18268. [Abstract] [Full Text] [PDF]