The gene cluster directing O-antigen biosynthesis in Yersinia enterocolitica serotype O:8: identification of the genes for mannose and galactose biosynthesis and the gene for the O-antigen polymerase

Abstract

The rfb gene cluster of Yersinia entemcolitica serotype 0:8 (Ye08) strain 8081 -c was cloned by cosmid cloning. Restriction mapping, deletion analysis and transposon mutagenesis showed that about 19 kb of the cloned DNA is Kiinamyllynkatu 13, 20520 Turku, Finland essential for the synthesis and expression of the Ye08 0-side-chain in Escherichia coli. Deletion analysis generated a derivative that expressed semirough LPS, a phenotype typical of an rfc mutant lacking the 0-antigen polymerase. The deletions and transcomplementation experiments allowed localization of the rfc gene to the 3'-end of the rfb gene cluster. The deduced Ye08 Rfc did not share significant amino acid sequence similarity with any other protein, but its amino acid composition and hydrophobicity profile are similar to those of identified Rfc proteins. In addition, the codon usage of the rfc gene is similar to other rfc genes. Nucleotide sequence analysis identified three other genes upstream of rfc. Two of the gene products showed 60-70°/0 identity to the RfbM and RfbK proteins that are biosynthetic enzymes for the GDPmannose pathway of enterobacteria. The third gene product was about 50-80 O/ o identical to the bacterial GalE protein, UDPglucose bepimerase, which catalyses the epimerization of UDPglucose to UDPgalactose. Since mannose and galactose are both present in the Ye08 0-antigen repeat unit, the above three genes are likely to belong to the rib gene cluster. A gene similar to the gsk gene downstream of rfc, and genes similar to adk and hemH upstream of the rfb gene cluster, were recognized. Thus the gene cluster of Ye08 is located between the ad&-hemH and gsk loci, and the order is adk-hemH-dbfc-gsk in the chromosome. Also in other Yersinia spp., the locus downstream of the hemH gene is occupied by gene clusters associated with LPS biosynthesis.

Evolutionary Dynamics of the Yersinia enterocolitica Complex

“…It has been shown before that the O:8 encoding locus is in the same genomic context as the outer core operon of serotypes O:3 and O:9 (Zhang, Toivanen and Skurnik, 1996;Skurnik and Bengoechea, 2003;Figure 4.12). The outer core is located beween gsk (inosine-guanosine kinase, YE3069) and hemH (ferrochelatase, YE3088).…”

“…Apart from gsk and hemH, the UDP-glucose 4-epimerase galE has been conserved between the outer core and the O-antigen. The same locus also contains genes involved in the synthesis of the heteropolymeric lipopolysaccharides of Y. pseudotuberculosis and Y. pestis, although five genes are inactivated in the latter (Zhang, Toivanen and Skurnik, 1996;Skurnik and Bengoechea, 2003 The absence of the outer core in the genome data for the other environmental…”

“…In O:3 and O:9 O-antigens the chain consists of unbranched homopolymers (Zhang, Toivanen and Skurnik, 1996;Skurnik and Bengoechea, 2003). The outer core of O:3 and O:9 serotype strains consists of a branched heteropolymer (Reeves, et al, 1996b;Skurnik, et al, 1999).…”

Localization and molecular characterization of putative O antigen gene clusters of Providencia species

“…In Pseudomonas aeruginosa, the OGC is located between himD and tyrB (Raymond et al, 2002), in Vibrio cholerae between gmhD and rjg (Sozhamannan et al, 1999), in Yersinia spp. between hemH and gsk (Pacinelli et al, 2002;Zhang et al, 1996), and in Proteus mirabilis between cpxA and secB (Wang et al, 2010). However, in Providencia, no OGC has been found and characterized so far.…”

“…Zhang et al, 1996), and in Proteus mirabilis between cpxA and secB (Wang et al, 2010). However, in Providencia, no OGC has been found and characterized so far.…”