N-Acetylmuramic Acid as Capping Element of α-D-Fucose-containing S-layer Glycoprotein Glycans from Geobacillus tepidamans GS5–97T

Kählig, Hanspeter; Kolarich, Daniel; Zayni, Sonja; Scheberl, Andrea; Kosma, Paul; Schäffer, Christina; Messner, Paul

doi:10.1074/jbc.m501724200

Cited by 25 publications

(29 citation statements)

References 39 publications

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“…The methyltransferase domain of ORFG106 is probably responsible for this terminal modification and may be involved in polysaccharide chain-length regulation analogous to WbdD O8 and WbdD O9a (36). Recently, the S-layer glycan of Geobacillus tepidamans GS5-97T was found to terminate in an N-acetylmuramic acid moiety (69). Thus, the significance of the results reported for the polymannan O8 and O9a export system may extend beyond O-PS.…”

Section: Discussionmentioning

confidence: 92%

The C-terminal Domain of the Nucleotide-binding Domain Protein Wzt Determines Substrate Specificity in the ATP-binding Cassette Transporter for the Lipopolysaccharide O-antigens in Escherichia coli Serotypes O8 and O9a

Cuthbertson¹,

Powers²,

Whitfield

2005

Journal of Biological Chemistry

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The polymannan O-antigenic polysaccharides (O-PSs) of Escherichia coli O8 and O9a are synthesized via an ATP-binding cassette (ABC) transporter-dependent pathway. The group 2 capsular polysaccharides of E. coli serve as prototypes for polysaccharide synthesis and export via this pathway. Here, we show that there are some fundamental differences between the ABC transporter-dependent pathway for O-PS biosynthesis and the capsular polysaccharide paradigm. In the capsule system, mutants lacking the ABC transporter are viable, and membranes isolated from these strains are no longer able to synthesize polymer using an endogenous acceptor. In contrast, E. coli strains carrying mutations in the membrane component (Wzm) and/or the nucleotide-binding component (Wzt) of the O8 and O9a polymannan transporters are nonviable under conditions permissive to O-PS biosynthesis and take on an aberrant elongated cell morphology. Whereas the ABC transporters for capsular polysaccharides with different structures are functionally interchangeable, the O8 and O9a exporters are specific for their cognate polymannan substrates. The E. coli O8 and O9a Wzt proteins contain a C-terminal domain not present in the corresponding nucleotide-binding protein (KpsT) from the capsule exporter. Whereas the Wzm components are functionally interchangeable, albeit with reduced efficiency, the Wzt components are not, indicating a specific role for Wzt in substrate specificity. Chimeric Wzt proteins were constructed in order to localize the region involved in substrate specificity to the C-terminal domain. ATP-binding cassette (ABC)1 transporters, or traffic ATPases, are responsible for the import and export of a variety of molecules across membranes. In the Escherichia coli K-12 genome, 79 known or putative ABC transporters have been identified indicating the importance of this protein superfamily in cellular physiology (1). The prototypical transporter consists of four domains (two transmembrane domains (TMDs) and two nucleotide-binding domains (NBDs)), which may be organized in a variety of ways (2). The TMD components between different systems share low sequence similarity and contain a variable number of transmembrane segments, whereas the NBD proteins of different systems share a higher overall sequence similarity as a result of the conserved sequence motifs required for ATP hydrolysis (3). X-ray structures are available for a number of NBD proteins as well as four complete transporters (reviewed in Refs. 4 -7). The structures reveal a common organization of the NBD monomers into two subdomains: a RecAlike domain and a helical domain not found in other ATPhydrolyzing proteins (4). The ATP-binding site is found along the NBD dimer interface and is made up of conserved residues from each monomer, namely the Walker A motif from one monomer and the ABC signature motif from the other monomer (8). Because the ABC transporter TMD components differ greatly between systems, whereas the NBD components remain highly similar, ABC transporters have been said to...

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Section: Discussionmentioning

confidence: 92%

The C-terminal Domain of the Nucleotide-binding Domain Protein Wzt Determines Substrate Specificity in the ATP-binding Cassette Transporter for the Lipopolysaccharide O-antigens in Escherichia coli Serotypes O8 and O9a

Cuthbertson¹,

Powers²,

Whitfield

2005

Journal of Biological Chemistry

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“…The Klebsiella O12 antigen contains a terminal 3-deoxy-D-manno-octulosonic acid residue, a sugar predominantly present in the LPS core (15). An S-layer glycan from Geobacillus tepidamans is terminated with both N-acetylglucosamine and N-acetylmuramic acid (37). N-Acetylmuramic acid was generally thought to be unique to peptidoglycan in the bacterial cell wall.…”

Section: Discussionmentioning

confidence: 99%

In Vitro Reconstruction of the Chain Termination Reaction in Biosynthesis of the Escherichia coli O9a O-Polysaccharide

Clarke

Richards

Greenfield

et al. 2011

Journal of Biological Chemistry

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Background: WbdD is a chain-length regulator that modifies the non-reducing terminus of the Escherichia coli O9a glycan. Results: WbdD phosphorylates and methylates a synthetic O9a repeating unit acceptor. Conclusion:The acceptor is modified with a terminal methyl phosphate. Significance: Determining the terminal structure is crucial to understanding the quality control processes of chain-length regulation and export of this prototypical glycan.

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“…Deletion of this enzyme resulted not only in an irregular thylakoid organization, but also in an altered cell wall and loss of the S-layer in the ⌬sll1213 mutant. Fucose has been identified as a building block in the Slayer glycan of Geobacillus tepidamans (40) and may be important for proper anchoring of the S-layer protein to the underlying lipopolysaccharide. Changes in composition of the lipopolysaccharide layer or (if present) the S-layer glycan frequently lead to a loss of the glycocalyx (41)(42)(43).…”

Section: Discussionmentioning

confidence: 99%

The sll1951 Gene Encodes the Surface Layer Protein of Synechocystis sp. Strain PCC 6803

Trautner

Vermaas

2013

Journal of Bacteriology

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Sll1951 is the surface layer (S-layer) protein of the cyanobacterium Synechocystis sp. strain PCC 6803. This large, hemolysin-like protein was found in the supernatant of a strain that was deficient in S-layer attachment. An sll1951 deletion mutation was introduced into Synechocystis and was easily segregated to homozygosity under laboratory conditions. By thin-section and negativestain transmission electron microscopy, a ϳ30-nm-wide S-layer lattice covering the cell surface was readily visible in wild-type cells but was absent in the ⌬sll1951 strain. Instead, the ⌬sll1951 strain displayed a smooth lipopolysaccharide surface as its most peripheral layer. In the presence of chaotropic agents, the wild type released a large (>150-kDa) protein into the medium that was identified as Sll1951 by mass spectrometry of trypsin fragments; this protein was missing in the ⌬sll1951 strain. In addition, Sll1951 was prominent in crude extracts of the wild type, indicating that it is an abundant protein. The carotenoid composition of the cell wall fraction of the ⌬sll1951 strain was similar to that of the wild type, suggesting that the S-layer does not contribute to carotenoid binding. Although the photoautotrophic growth rate of the ⌬sll1951 strain was similar to that of the wild-type strain, the viability of the ⌬sll1951 strain was reduced upon exposure to lysozyme treatment and hypo-osmotic stress, indicating a contribution of the S-layer to the integrity of the Synechocystis cell wall. This work identifies the S-layer protein in Synechocystis and shows that, at least under laboratory conditions, this very abundant, large protein has a supportive but not a critical role in the function of the cyanobacterium.

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N-Acetylmuramic Acid as Capping Element of α-D-Fucose-containing S-layer Glycoprotein Glycans from Geobacillus tepidamans GS5–97T

Cited by 25 publications

References 39 publications

The C-terminal Domain of the Nucleotide-binding Domain Protein Wzt Determines Substrate Specificity in the ATP-binding Cassette Transporter for the Lipopolysaccharide O-antigens in Escherichia coli Serotypes O8 and O9a

The C-terminal Domain of the Nucleotide-binding Domain Protein Wzt Determines Substrate Specificity in the ATP-binding Cassette Transporter for the Lipopolysaccharide O-antigens in Escherichia coli Serotypes O8 and O9a

In Vitro Reconstruction of the Chain Termination Reaction in Biosynthesis of the Escherichia coli O9a O-Polysaccharide

The sll1951 Gene Encodes the Surface Layer Protein of Synechocystis sp. Strain PCC 6803

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