Functional Characterization of WaaL, a Ligase Associated with Linking O-Antigen Polysaccharide to the Core of
            <i>Pseudomonas aeruginosa</i>
            Lipopolysaccharide

Abeyrathne, Priyanka D.; Daniels, Craig; Poon, Karen; Matewish, Mauricia J.; Lam, Joseph S.

doi:10.1128/jb.187.9.3002-3012.2005

Cited by 134 publications

(127 citation statements)

References 64 publications

(61 reference statements)

Supporting

Mentioning

119

Contrasting

Order By: Relevance

“…aeruginosa plays a role in flagellar biogenesis in this species (Abeyrathne et al, 2005). However, this is not the case in A. hydrophila galU mutants because the flagellar biogenesis is unaffected.…”

Section: Discussionmentioning

confidence: 73%

Mesophilic Aeromonas UDP-glucose pyrophosphorylase (GalU) mutants show two types of lipopolysaccharide structures and reduced virulence

et al. 2007

View full text Add to dashboard Cite

A mutation in galU that causes the lack of O34-antigen lipopolysaccharide (LPS) in Aeromonas hydrophila strain AH-3 was identified. It was proved that A. hydrophila GalU is a UDP-glucose pyrophosphorylase responsible for synthesis of UDP-glucose from glucose 1-phosphate and UTP. The galU mutant from this strain showed two types of LPS structures, represented by two bands on LPS gels. The first one (slow-migrating band in gels) corresponds to a rough strain having the complete core, with two significant differences: it lacks the terminal galactose residue from the LPS-core and 4-amino-4-deoxyarabinose residues from phosphate groups in lipid A. The second one (fast-migrating band in gels) corresponds to a deeply truncated structure with the LPS-core restricted to one 3-deoxy-D-manno-oct-2-ulosonic acid (Kdo) and three L-glycero-Dmanno-heptose residues. galU mutants in several motile mesophilic Aeromonas strains from serotypes O1, O2, O11, O18, O21 and O44 were also devoid of the O-antigen LPS. The galU mutation reduced to less than 1 % the survival of these Aeromonas strains in serum, decreased the ability of these strains to adhere and reduced by 1.5 or 2 log units the virulence of Aeromonas serotype O34 strains in a septicaemia model in either fish or mice. All the changes observed in the galU mutants were rescued by the introduction of the corresponding single wild-type gene.

show abstract

Section: Discussionmentioning

confidence: 73%

Mesophilic Aeromonas UDP-glucose pyrophosphorylase (GalU) mutants show two types of lipopolysaccharide structures and reduced virulence

et al. 2007

View full text Add to dashboard Cite

show abstract

“…Thus, these results are consistent with the normal production of O-antigen precursors that cannot be attached to the core OS because of a wapR knockout. Our group has previously shown that this type of undecaprenol phosphate-linked O-antigens is sensitive to phenol (1,4,20), and when LPS from the wapR mutant was prepared by the hot aqueous phenol method of Westphal and Jann (31) rather than the fast method of Hitchcock and Brown (10), A-band and B-band O antigens were not detected by either SDS-PAGE or Western immunoblotting (data not shown).…”

Section: Discussionmentioning

confidence: 99%

Functional Characterization of MigA and WapR: Putative Rhamnosyltransferases Involved in Outer Core Oligosaccharide Biosynthesis ofPseudomonas aeruginosa

et al. 2008

Self Cite

View full text Add to dashboard Cite

Pseudomonas aeruginosa lipopolysaccharide (LPS) contains two glycoforms of core oligosaccharide (OS); one form is capped with O antigen through an ␣-1,3-linked L-rhamnose (L-Rha), while the other is uncapped and contains an ␣-1,6-linked L-Rha. Two genes in strain PAO1, wapR (PA5000) and migA (PA0705), encode putative glycosyltransferases associated with core biosynthesis. We propose that WapR and MigA are the rhamnosyltransferases responsible for the two linkages of L-Rha to the core. Knockout mutants with mutations in both genes were generated. The wapR mutant produced LPS lacking O antigen, and addition of wapR in trans complemented this defect. The migA mutant produced LPS with a truncated outer core and showed no reactivity to outer core-specific monoclonal antibody (MAb) 5C101. Complementation of this mutant with migA restored reactivity of the LPS to MAb 5C101. Interestingly, LPS from the complemented migA strain was not reactive to MAb 18-19 (specific for the core-plus-one O repeat). This was due to overexpression of MigA in the complemented strain that caused an increase in the proportion of the uncapped core OS, thereby decreasing the amount of the core-plus-one O repeat, indicating that MigA has a regulatory role. The structures of LPS from both mutants were elucidated using nuclear magnetic resonance spectroscopy and mass spectrometry. The capped core of the wapR mutant was found to be truncated and lacked ␣-1,3-L-Rha. In contrast, uncapped core OS from the migA mutant lacked ␣-1,6-L-Rha. These results provide evidence that WapR is the ␣-1,3-rhamnosyltransferase, while MigA is the ␣-1,6-rhamnosyltransferase.Pseudomonas aeruginosa is an important gram-negative opportunistic pathogen capable of producing virulence determinants, including secreted toxins and enzymes, as well as cell surface structures, such as adhesins, flagella, and lipopolysaccharide (LPS) (18). LPS is a glycolipid composed of three distinct regions: lipid A, core oligosaccharide, and O antigen. P. aeruginosa produces two forms of O antigen, which are the homopolymeric A band and the heteropolymeric B band (15,21). The gene clusters associated with the biosynthesis of Aband and B-band O antigen in P. aeruginosa have been well characterized (for reviews, see references 18 and 22). In contrast, relatively little is known about the biosynthesis of the core oligosaccharide (OS).The P. aeruginosa core OS, like that of other gram-negative bacteria, can be conceptually subdivided into the inner core and the outer core (Fig. 1). The inner core is highly conserved and contains two L-glycero-D-manno-heptose (Hep) residues and two 3-deoxy-D-manno-octulosonic acid (Kdo) residues; the genetics and enzymology of this region are the best understood to date (14). The outer core is composed of four D-glucose (D-Glc) residues, one L-rhamnose (L-Rha) residue, and one N-(L-alanyl)-D-galactosamine residue, and its biosynthesis is not fully understood. The structures of the core OS of several different serotypes, clinical isolates, and rough mutants of P. a...

show abstract

“…These repeat units are then sequentially assembled to a growing chain by the polymerase Wzy, and the length of the growing polysaccharide is determined by the polymerase co-polymerase (PCP) Wzz [17]. The complete periplasmic O-antigen chain is attached by the ligase WaaL to the lipid A-core moiety anchored to the outer leaflet of the IM [1], forming smooth-LPS.…”

Section: Introductionmentioning

confidence: 99%

Characterization of interactions between LPS transport proteins of the Lpt system

Bowyer

Baardsnes

Ajamian

et al. 2011

Biochemical and Biophysical Research Communications

View full text Add to dashboard Cite

The lipopolysaccharide transport system (Lpt) in Gram-negative bacteria is responsible for transporting lipopolysaccharide (LPS) from the cytoplasmic surface of the inner membrane, where it is assembled, across the inner membrane, periplasm and outer membrane, to the surface where it is then inserted in the outer leaflet of the asymmetric lipid bilayer. The Lpt system consists of seven known LPS transport proteins (LptA-G) spanning from the cytoplasm to the cell surface. We have shown that the periplasmic component, LptA is able to form a stable complex with the inner membrane anchored LptC but does not interact with the outer membrane anchored LptE. This suggests that the LptC component of the LptBFGC complex may act as a dock for LptA, allowing it to bind LPS after it has been assembled at the inner membrane. That no interaction between LptA and LptE has been observed supports the theory that LptA binds LptD in the LptDE homodimeric complex at the outer membrane.Crown

show abstract

Functional Characterization of WaaL, a Ligase Associated with Linking O-Antigen Polysaccharide to the Core of Pseudomonas aeruginosa Lipopolysaccharide

Cited by 134 publications

References 64 publications

Mesophilic Aeromonas UDP-glucose pyrophosphorylase (GalU) mutants show two types of lipopolysaccharide structures and reduced virulence

Mesophilic Aeromonas UDP-glucose pyrophosphorylase (GalU) mutants show two types of lipopolysaccharide structures and reduced virulence

Functional Characterization of MigA and WapR: Putative Rhamnosyltransferases Involved in Outer Core Oligosaccharide Biosynthesis ofPseudomonas aeruginosa

Characterization of interactions between LPS transport proteins of the Lpt system

Contact Info

Product

Resources

About