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

J. Biol. Chem., Vol. 281, Issue 18, 12879-12887, May 5, 2006

This Article Full Text Full Text (PDF) All Versions of this Article: 281/18/12879    most recent M513865200v1 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 Kanjilal-Kolar, S. Articles by Raetz, C. R. H. Search for Related Content PubMed PubMed Citation Articles by Kanjilal-Kolar, S. Articles by Raetz, C. R. H. Social Bookmarking                      What's this?

Dodecaprenyl Phosphate-Galacturonic Acid as a Donor Substrate for Lipopolysaccharide Core Glycosylation in Rhizobium leguminosarum^*

From the Department of Biochemistry, Duke University Medical Center, Durham, North Carolina 27710

The lipid A and inner core regions of Rhizobium leguminosarum lipopolysaccharide contain four galacturonic acid (GalA) residues. Two are attached to the outer unit of the 3-deoxy-D-manno-octulosonic acid (Kdo) disaccharide, one to the mannose residue, and one to the 4'-position of lipid A. The enzymes RgtA and RgtB, described in the accompanying article, catalyze GalA transfer to the Kdo residue, whereas RgtC is responsible for modification of the core mannose unit. Heterologous expression of RgtA in Sinorhizhobium meliloti 1021, a strain that normally lacks GalA modifications on its Kdo disaccharide, resulted in detectable GalA transferase activity in isolated membrane preparations, suggesting that the appropriate GalA donor substrate is available in S. meliloti membranes. In contrast, heterologous expression of RgtA in Escherichia coli yielded inactive membranes. However, RgtA activity was detectable in the E. coli system when total lipids from R. leguminosarum 3841 or S. meliloti 1021 were added. We have now purified and characterized dodecaprenyl (C60) phosphate-GalA as a minor novel lipid of R. leguminosarum 3841 and S. meliloti. This substance is stable to mild base hydrolysis and was purified by DEAE-cellulose column chromatography. Its structure was established by a combination of electrospray ionization mass spectrometry and gas-liquid chromatography. Purified dodecaprenyl phosphate-GalA supports the efficient transfer of GalA to Kdo₂-1-dephospho-lipid IV_A by membranes of E. coli cells expressing RgtA, RgtB, and RgtC. The identification of a polyisoprene phosphate-GalA donor substrate suggests that the active site of RgtA faces the periplasmic side of the inner membrane. This work represents the first definitive characterization of a lipid-linked GalA derivative with the proposed structure dodecaprenyl phosphate--D-GalA.

Received for publication, December 29, 2005 , and in revised form, February 22, 2006.

^* This work was supported by National Institutes of Health Grant GM-51796 (to C. R. H. R.). 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.

¹ To whom correspondence should be addressed: Duke University Medical Center, Durham, NC, 27710. Tel.: 919-684-5326; Fax: 919-684-8885; E-mail, raetz{at}biochem.duke.edu.

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

This article has been cited by other articles:

				K. A. Grabinska, S. K. Ghosh, Z. Guan, J. Cui, C. R. H. Raetz, P. W. Robbins, and J. Samuelson Dolichyl-Phosphate-Glucose Is Used To Make O-Glycans on Glycoproteins of Trichomonas vaginalis Eukaryot. Cell, August 1, 2008; 7(8): 1344 - 1351. [Abstract] [Full Text] [PDF]

				A. Yan, Z. Guan, and C. R. H. Raetz An Undecaprenyl Phosphate-Aminoarabinose Flippase Required for Polymyxin Resistance in Escherichia coli J. Biol. Chem., December 7, 2007; 282(49): 36077 - 36089. [Abstract] [Full Text] [PDF]

				S. Kanjilal-Kolar, S. S. Basu, M. I. Kanipes, Z. Guan, T. A. Garrett, and C. R. H. Raetz Expression Cloning of Three Rhizobium leguminosarum Lipopolysaccharide Core Galacturonosyltransferases J. Biol. Chem., May 5, 2006; 281(18): 12865 - 12878. [Abstract] [Full Text] [PDF]