A GENE (WBBL) FROM SERRATIA-MARCESCENS N28B (O4) COMPLEMENTS THE RFB-50 MUTATION OF ESCHERICHIA-COLI K-12 DERIVATIVES

A cosmid-based genomic library of Serratia marcescens N28b was introdu ced into Escherichia coli DH5 alpha, and clones were screened for seru m resistance. One clone was found resistant to serum, to bacteriocin 2 8b, and to bacteriophages TuIa and TuIb. This clone also showed O anti gen in its lipopolysaccharide. Subcloning and sequencing experiments s howed that a 2,124-bp DNA fragment containing the rmlD and wbbL genes was responsible for the observed phenotypes. On the basis of amino aci d similarity, we suggest that the 288-residue RmlD protein is a dTDP-L -rhamnose synthase. Plasmid pJT102, containing only the wbbL, gene, wa s able to induce O16-antigen production and serum resistance in E. col i DH5 alpha. These results suggest that the 282-residue WbbL protein i s a rhamnosyltransferase able to complement the rfb-50 mutation in E. coli K-12 derivatives, despite the low level of amino acid identity be tween WbbL and the E. coli rhamnosyltransferase (24.80%). S. marcescen s N28b rmlD and wbbL mutants were constructed by mobilization of suici de plasmids containing a portion of rmlD or wbbL. These insertion muta nts were unable to produce O antigen; since strain N28b produces O4 an tigen, these results suggest that both genes are involved in O 4-antig en biosynthesis.