Cloning and characterization of the Escherichia coli K-12 rfa-2 (rfaC) gene, a gene required for lipopolysaccharide inner core synthesis.

摘要: A genetically defined mutation, designated rfa-2, results in altered lipopolysaccharide (LPS) biosynthesis. rfa-2 mutants produce a core-defective LPS that contains lipid and single sugar moiety, 2-keto-3-deoxyoctulosonic acid, the core region. Such or deep-rough (R) mutant structures were previously chemotype Re. Phenotypically, exhibit increased permeability to number of hydrophilic hydrophobic agents. By restriction analyses complementation studies, we clearly gene on 1,056-bp AluI-DraI fragment. The flanking rfa locus regions completely sequenced. Additionally, location physical map Escherichia coli chromosome was determined. encodes 36,000-dalton polypeptide an vivo expression system. N-terminal analysis purified product confirmed first 24 amino acid residues as deduced from nucleotide sequence coding interspecies complementation, Salmonella typhimurium rfaC (LPS Re) is transformed with E. rfa-2+ gene, transformant characterized by wild-type sensitivity novobiocin (i.e., uninhibited growth at 600 micrograms per ml) restoration ability synthesize structures. On basis identity significant similarity its recently (D. M. Sirisena, K. A. Brozek, P. R. MacLachlan, Sanderson, C. H. Raetz, J. Biol. Chem. 267:18874-18884, 1992), S. (heptosyltransferase 1), K-12 will be rfaC.