Smith-degradative studies on the polysaccharide portion of A-band lipopolysaccharide from a mutant (AK1401) of <i>Pseudomonas</i> <i>aeruginosa</i> strain PAO1

Arsenault, Todd L.; MacLean, David B.; Zou, W.; Szarek, Walter A.

doi:10.1139/v94-172

Cited by 7 publications

(2 citation statements)

References 6 publications

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“…NMR spectroscopic data reported here for the d ‐rhamnan and SD1 from S. maltophilia are comparable with those obtained in other studies, allowing for variations in recording parameters and the tentative assignments for some signals, and the value of the specific rotation of the O7 antigen was also similar to values obtained for other rhamnans [20, 21, 24]. The formation of by‐products identical to SD2 and SD3 was also observed during Smith degradation of the common antigen of P. aeruginosa [26]. It may also be noted that a monoclonal antibody (E87) specific to the common antigen of P. aeruginosa recognizes some strains of S. maltophilia [27], although the surface polysaccharides of these strains were not characterized, and d ‐rhamnose residues occur in different O antigens from S. maltophilia ([19, 28], and authors' unpublished results).…”

Section: Resultssupporting

confidence: 87%

The O7 antigen of Stenotrophomonas maltophilia is a linear d-rhamnan with a trisaccharide repeating unit that is also present in polymers from some Pseudomonas and Burkholderia species

Winn

Wilkinson

1998

FEMS Microbiology Letters

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The O antigen polymer recovered from the reference strain for Stenotrophomonas (Xanthomonas or Pseudomonas) maltophilia serogroup O7, after mild acid hydrolysis of the lipopolysaccharide, was constructed from D-rhamnose. By means of chemical degradations and NMR studies, the repeating unit of the polymer was shown to be a linear trisaccharide with the structure -->2)-alpha-D-Rhap-(1-->3)-alpha-D-Rhap-(1-->3)-alpha-D-R hap-(1-->. The same repeating unit is present in the common antigen of Pseudomonas aeruginosa and in O antigens from some pathovars of Pseudomonas syringae and a strain of Burkholderia (Pseudomonas) cepacia.

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Section: Resultssupporting

confidence: 87%

The O7 antigen of Stenotrophomonas maltophilia is a linear d-rhamnan with a trisaccharide repeating unit that is also present in polymers from some Pseudomonas and Burkholderia species

Winn

Wilkinson

1998

FEMS Microbiology Letters

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“…2-D-Rha␣1-3-D-Rha␣1-3] (7,8). D-Rha is a rare sugar in bacterial polysaccharides and is not found in human glycoconjugates.…”

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confidence: 99%

Biosynthesis of the Common Polysaccharide Antigen of Pseudomonas aeruginosa PAO1: Characterization and Role of GDP- d -Rhamnose:GlcNAc/GalNAc-Diphosphate-Lipid α1,3- d -Rhamnosyltransferase WbpZ

et al. 2015

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The opportunistic pathogen Pseudomonas aeruginosa produces two major cell surface lipopolysaccharides, characterized by distinct O antigens, called common polysaccharide antigen (CPA) and O-specific antigen (OSA). CPA contains a polymer of Drhamnose (D-Rha) in ␣1-2 and ␣1-3 linkages. Three putative glycosyltransferase genes, wbpX, wbpY, and wbpZ, are part of the CPA biosynthesis cluster. To characterize the enzymatic function of the wbpZ gene product, we chemically synthesized the donor substrate GDP-D-Rha and enzymatically synthesized GDP-D-[ 3 H]Rha. Using nuclear magnetic resonance (NMR) spectroscopy, we showed that WbpZ transferred one D-Rha residue from GDP-D-Rha in ␣1-3 linkage to both GlcNAc-and GalNAcdiphosphate-lipid acceptor substrates. WbpZ is also capable of transferring D-mannose (D-Man) to these acceptors. Therefore, WbpZ has a relaxed specificity with respect to both acceptor and donor substrates. The diphosphate group of the acceptor, however, is required for activity. WbpZ does not require divalent metal ion for activity and exhibits an unusually high pH optimum of 9. WbpZ from PAO1 is therefore a GDP-D-Rha:GlcNAc/GalNAc-diphosphate-lipid ␣1,3-D-rhamnosyltransferase that has significant activity of GDP-D-Man:GlcNAc/GalNAc-diphosphate-lipid ␣1,3-D-mannosyltransferase. We used site-directed mutagenesis to replace the Asp residues of the two DXD motifs with Ala. Neither of the mutant constructs of wbpZ (D172A or D254A) could be used to rescue CPA biosynthesis in the ⌬wbpZ knockout mutant in a complementation assay. This suggested that D172 and D254 are essential for WbpZ function. This work is the first detailed characterization study of a D-Rha-transferase and a critical step in the development of CPA synthesis inhibitors. IMPORTANCEThis is the first characterization of a D-rhamnosyltransferase and shows that it is essential in Pseudomonas aeruginosa for the synthesis of the common polysaccharide antigen. P seudomonas aeruginosa is a Gram-negative bacterium that is ubiquitous in the environment and is an opportunistic pathogen that can cause life-threatening infections in humans whose defenses are compromised, e.g., those with immune deficiency, burn wounds, cancer, or cystic fibrosis (CF). Specific epidemic strains have been shown to cause local outbreaks and hospital-associated infections (1). In the airways of CF patients, P. aeruginosa thrives, adopting a biofilm lifestyle, and often becomes resistant to antibiotic treatment. Therefore, it is important to understand the mechanisms by which P. aeruginosa synthesizes its virulence factor, in order to develop new antibacterial strategies.Lipopolysaccharides (LPS) on the outer membrane of P. aeruginosa are required for its survival against host defense mechanisms; hence, LPS is one of the major virulence factors (2-4). These bacteria are unusual in that they simultaneously synthesize two distinct forms of LPS differing in the O-antigen structures (5). Each of these O antigens is synthesized by a different pathway.

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A convenient synthesis of GDP-d-rhamnose: The donor substrate for d-rhamnosyltransferase WbpZ from Pseudomonas aeruginosa PAO1

Wang

Tanaka

Hindsgaul

et al. 2013

Bioorganic & Medicinal Chemistry Letters

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Smith-degradative studies on the polysaccharide portion of A-band lipopolysaccharide from a mutant (AK1401) of Pseudomonas aeruginosa strain PAO1

Abstract: , 1376 (1994). Hepta-O-acetyl-O-a-~-Rhap-(1+3)-O-a-~-Rhap-(l+2)-glycerol

Cited by 7 publications

References 6 publications

The O7 antigen of Stenotrophomonas maltophilia is a linear d-rhamnan with a trisaccharide repeating unit that is also present in polymers from some Pseudomonas and Burkholderia species

The O7 antigen of Stenotrophomonas maltophilia is a linear d-rhamnan with a trisaccharide repeating unit that is also present in polymers from some Pseudomonas and Burkholderia species

Biosynthesis of the Common Polysaccharide Antigen of Pseudomonas aeruginosa PAO1: Characterization and Role of GDP- d -Rhamnose:GlcNAc/GalNAc-Diphosphate-Lipid α1,3- d -Rhamnosyltransferase WbpZ

A convenient synthesis of GDP-d-rhamnose: The donor substrate for d-rhamnosyltransferase WbpZ from Pseudomonas aeruginosa PAO1

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