Role of Rfe and RfbF in the initiation of biosynthesis of D-galactan I, the lipopolysaccharide O antigen from Klebsiella pneumoniae serotype O1

Abstract

The 6.6-kb rfb gene cluster from Klebsiella pneumoniae serotype O1 (rfb KpO1 Rfe is an N-acetylglucosamine (GlcpNAc)-1-phosphate transferase which forms lipid I (undecaprenyl pyrophosphoryl GlcpNAc) during the biosynthesis of the cell surface polysaccharide known as enterobacterial common antigen (37). In members of the Enterobacteriaceae, Rfe can also be involved in both pathways for LPS O antigen synthesis. In the Rfc-dependent pathway, Rfe transfers GlcpNAc residues to a lipid intermediate to initiate the formation of each O polysaccharide repeat unit (1,29,58,72). rfb gene products then complete the various O antigen repeat units, and after polymerization, GlcpNAc residues provide the first monomer in each repeat unit. A different role is played by Rfe in the biosynthesis of some homopolymeric O polysaccharides which, like D-galactan I, lack GlcpNAc in their repeating unit structure. In E. coli O8 and O9, Rfe primes the synthesis of the mannose homopolymer O polysaccharide by forming an undecaprenyl pyrophos-

A Membrane-located Glycosyltransferase Complex Required for Biosynthesis of the d-Galactan I Lipopolysaccharide O Antigen in Klebsiella pneumoniae

“…The predicted N-terminal domain (WbbO N ) could not complement the ⌬wbbO mutation as expected (data not shown). Therefore, when expressed from a plasmid, WbbO does not require the N terminus for galactosyltransferase function and the reported ␣-galactopyranosyl-and ␤-galactofuranosyltransferase activity (14,20) of the enzyme is confined to the C-terminal domain of WbbO.…”

“…From both in vivo and in vitro experiments, WbbO is proposed (11). The adaptor region consists of two galactose residues believed to be added to und-PP-GlcNAc by WbbO (14,20). The precise glycosyltransferase activities required for extension of the D-galactan I domain remain undetermined.…”

“…to add the first two sugars (Galp-Galf) to the und-PP-GlcNAc acceptor (14,20), forming the adaptor region of the O-PS (Fig. 1).…”

“…It is responsible for forming the adaptor region (14,20). Its activity is followed by WbbM (20), implicating this GT-8 enzyme as an ␣1,3-Galp-transferase.…”

“…The acceptor is synthesized by the UDP-N-acetylglucosamine:Und-P GlcNAc-1-P transferase, WecA (10 -13). This requirement has been established for D-galactan-I (14). O-PS is elongated on this acceptor at the cytoplasmic face of the inner membrane by dedicated O-antigen-specific glycosyltransferases.…”