JBC Oz Biosciences

HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
QUICK SEARCH:   [advanced]


     

Originally published In Press as doi:10.1074/jbc.M004191200 on August 7, 2000

J. Biol. Chem., Vol. 275, Issue 43, 33252-33259, October 27, 2000
This Article
Right arrow Full Text
Right arrow Full Text (PDF)
Right arrow An addition or correction has been published
Right arrow All Versions of this Article:
275/43/33252    most recent
M004191200v1
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow reprints & permissions
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Zhao, X.
Right arrow Articles by Lam, J. S.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Zhao, X.
Right arrow Articles by Lam, J. S.
Social Bookmarking
Add to CiteULike   Add to Complore   Add to Connotea   Add to Del.icio.us   Add to Digg   Add to Reddit   Add to Technorati  
What's this?

WbpO, a UDP-N-acetyl-D-galactosamine Dehydrogenase from Pseudomonas aeruginosa Serotype O6*

Xin ZhaoDagger , Carole CreuzenetDagger §, Myriam BélangerDagger §, Emmanuel Egbosimba||, Jianjun Li**, and Joseph S. LamDagger Dagger Dagger

From the Dagger  Department of Microbiology, University of Guelph, Guelph, Ontario N1G 2W1 and the ** Institute for Biological Sciences, National Research Council, Ottawa, Ontario K1A OR6, Canada

WbpO is associated with B-band lipopolysaccharide biosynthesis in Pseudomonas aeruginosa serotype O6. This protein is thought to catalyze the enzymatic conversion of UDP-N-acetyl-D-galactosamine (UDP-GalNAc) to UDP-N-acetyl-D-galactosaminuronic acid (UDP-GalNAcA). WbpO was overexpressed with a C-terminal hexahistidine tag. The soluble form of expressed WbpO (WbpOSol) exhibited a secondary structure with 29.2% alpha -helix and 20.1% beta -strand. However, no enzymatic activity could be detected using either high performance anion exchange chromatography or capillary electrophoresis-mass spectrometry analysis. An insoluble form of expressed WbpO was purified in the presence of guanidine hydrochloride by immobilized metal ion affinity chromatography. After refolding, this preparation of WbpO (designated as WbpORf) exhibited stable secondary structure at pH 7.5 to 8.2, and it was enzymatically active. Capillary electrophoresis-mass spectrometry and tandem mass spectrometry analysis showed that WbpORf catalyzed the conversion of UDP-GalNAc to UDP-GalNAcA. 26 and 22% of the substrate could be converted to UDP-GalNAcA in the presence of NAD+ and NADP+ as the cofactors, respectively. The Km values of WbpORf for UDP-GalNAc, NAD+, and NADP+ were 7.79, 0.65, and 0.44 mM, respectively. WbpORf can also catalyze the conversion of UDP-GlcNAc to UDP-GlcNAcA. In conclusion, this is the first report of the overexpression, purification, and biochemical characterization of an NAD+/NADP+-dependent UDP-GalNAc dehydrogenase. Our results also complete the biosynthetic pathway for GalNAcA that is part of the O-antigen of P. aeruginosa serotype O6 lipopolysaccharide.

* This work was supported in part by Grant MT14796 (to J. S. L.) from the Medical Research Council of Canada.The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

§ Recipients of fellowships from the Canadian Cystic Fibrosis Foundation.

Current address: Dept. of Pathobiology, College of Verterinary Medicine, University of Florida, Gainesville, FL 32611-0880.

|| Recipient of the Canadian Commonwealth Scholarship.

Dagger Dagger To whom correspondence should be addressed: Dept. of Microbiology, University of Guelph, Guelph, Ontario N1G 2W1, Canada. Tel.: 519-824-4120, ext. 3823; Fax: 519-837-1820; E-mail: jlam@uoguelph.ca.

Copyright © 2000 by The American Society for Biochemistry and Molecular Biology, Inc.
Add to CiteULike CiteULike   Add to Complore Complore   Add to Connotea Connotea   Add to Del.icio.us Del.icio.us   Add to Digg Digg   Add to Reddit Reddit   Add to Technorati Technorati    What's this?


This article has been cited by other articles:


Home page
 J. Bacteriol.Home page
E. L. Westman, A. Preston, R. A. Field, and J. S. Lam
Biosynthesis of a Rare Di-N-Acetylated Sugar in the Lipopolysaccharides of both Pseudomonas aeruginosa and Bordetella pertussis Occurs via an Identical Scheme despite Different Gene Clusters
J. Bacteriol., September 15, 2008; 190(18): 6060 - 6069.
[Abstract] [Full Text] [PDF]

Home page
 J. Biol. Chem.Home page
W. L. Miller, M. J. Matewish, D. J. McNally, N. Ishiyama, E. M. Anderson, D. Brewer, J.-R. Brisson, A. M. Berghuis, and J. S. Lam
Flagellin Glycosylation in Pseudomonas aeruginosa PAK Requires the O-antigen Biosynthesis Enzyme WbpO
J. Biol. Chem., February 8, 2008; 283(6): 3507 - 3518.
[Abstract] [Full Text] [PDF]

Home page
 Mol Biol EvolHome page
M. M. Cunneen and P. R. Reeves
The Yersinia kristensenii O11 O-Antigen Gene Cluster was Acquired by Lateral Gene Transfer and Incorporated at a Novel Chromosomal Locus
Mol. Biol. Evol., June 1, 2007; 24(6): 1355 - 1365.
[Abstract] [Full Text] [PDF]

Home page
 Infect. Immun.Home page
N. Y. Park, J. H. Lee, M. W. Kim, H. G. Jeong, B. C. Lee, T. S. Kim, and S. H. Choi
Identification of the Vibrio vulnificus wbpP Gene and Evaluation of Its Role in Virulence
Infect. Immun., January 1, 2006; 74(1): 721 - 728.
[Abstract] [Full Text] [PDF]

Home page
 J Med MicrobiolHome page
J. L. Prior, R. G. Prior, P. G. Hitchen, H. Diaper, K. F. Griffin, H. R. Morris, A. Dell, and R. W. Titball
Characterization of the O antigen gene cluster and structural analysis of the O antigen of Francisella tularensis subsp. tularensis
J. Med. Microbiol., October 1, 2003; 52(10): 845 - 851.
[Abstract] [Full Text] [PDF]

Home page
 Infect. Immun.Home page
D.-Q. Xu, J. O. Cisar, N. Ambulos Jr., D. H. Burr, and D. J. Kopecko
Molecular Cloning and Characterization of Genes for Shigella sonnei Form I O Polysaccharide: Proposed Biosynthetic Pathway and Stable Expression in a Live Salmonella Vaccine Vector
Infect. Immun., August 1, 2002; 70(8): 4414 - 4423.
[Abstract] [Full Text] [PDF]

Home page
 J. Bacteriol.Home page
C. K. Raymond, E. H. Sims, A. Kas, D. H. Spencer, T. V. Kutyavin, R. G. Ivey, Y. Zhou, R. Kaul, J. B. Clendenning, and M. V. Olson
Genetic Variation at the O-Antigen Biosynthetic Locus in Pseudomonas aeruginosa
J. Bacteriol., July 1, 2002; 184(13): 3614 - 3622.
[Abstract] [Full Text] [PDF]


HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
 All ASBMB Journals   Molecular and Cellular Proteomics 
 Journal of Lipid Research   ASBMB Today 
Copyright © 2000 by the American Society for Biochemistry and Molecular Biology.