GENOMIC ORGANIZATION OF THE KLEBSIELLA-PNEUMONIAE CPS REGION RESPONSIBLE FOR SEROTYPE K2 CAPSULAR POLYSACCHARIDE SYNTHESIS IN THE VIRULENT-STRAIN CHEDID

The genomic organization of the chromosomal cps region that is respons ible for capsular polysaccharide synthesis in Klebsiella pneumoniae Ch edid (O1:K2) was investigated. Deletion analyses and Southern hybridiz ation studies suggested that the central region of the cloned 29-kh Ba mHI fragment is indispensable for K2 capsular polysaccharide synthesis . The 24,329-bp nucleotide sequence of the Klebsiella cps region was d etermined and deposited in the EMBL and GenBank databases through DDBJ and assigned accession number D21242. Nineteen possible open reading frames (ORFs) mere identified in the sequenced area. Among them, 13 OR Fs are very close to each other. Six of the 19 ORFs show considerable nucleotide sequence similarities to Salmonella typhimurium cpsG, cpsB, rfbP, and orf2.8, Escherichia coli gnd, and Haemophilus influenzae be xD, respectively. Moreover, the deduced amino acid sequence of the ORF 10 product demonstrated a highly hydrophobic profile and showed putati ve membrane topology similarity to Rickettsia prowazekii ATP/ADP trans locase. Nucleotide sequences similar to the o(54)-dependent promoter, as well as the usual -35 and -10 sequences, mere identified just upstr eam of ORF3, which is the first ORF in the polycistronic structure. Fu rthermore, a sequence (GGGCGGTAGCGT) found just downstream of the o(54 )-dependent promoter-like sequence was generally conserved among gene clusters implicated in cell surface polysaccharide synthesis, such as Salmonella rfb and viaB and E. coli kpsMT and rfaQPG. A possible trans criptional terminator with a hairpin loop structure found just downstr eam of ORF15 that is a homolog of E. coli gnd. K2 capsular polysacchar ide biosynthesis in E. coli K-12 depends on cpsB (mannose-1-phosphate guanyltransferase gene), and klebsiella cpsB, found in the downstream region of the polycistronic structure, was able to complement cpsB off . coil. Results of transposon insertion and promoter-cloning analyses were consistent with the results of nucleotide sequence analysis.