MOLECULAR CHARACTERIZATION AND ADHESIVE PROPERTIES OF CF29K, AN ADHESIN OF KLEBSIELLA-PNEUMONIAE STRAINS INVOLVED IN NOSOCOMIAL INFECTIONS

We previously described a CS31A-related protein, CF29K expressed by Kl ebsiella pneumoniae strains involved in nosocomial infections. In this study, we cloned and sequenced cf29A, the structural gene of the CF29 K protein, and showed that CF29K is an antigenic subtype of CS31A. The CF29K protein was found to be identical to the CS31A-L protein on the basis of biochemical and immunological properties. In contrast, the C S31A-H protein presented a different apparent molecular mass during so dium dodecyl sulfate-polyacrylamide gel electrophoresis, a different l imited degradation pattern with endopeptidase V8, and a specific confo rmational epitope. We cloned and sequenced the CS31A-L structural gene and confirmed that CF29K and CS31A-L are identical, but their major s ubunits differ from ClpG (the CS31A-H subtype major subunit) by one am ino acid at position 89 of the mature protein, which is a lysine in CF 29K instead of the asparagine in ClpG. Site-directed mutagenesis exper iments demonstrated that the biochemical and immunological differences between CS31A-H and CF29K or CS31A-L were dependent only on the amino acid at position 89 of the mature protein. To study the adhesive prop erties of CS31A-H and CF29K in the same Escherichia coli reference str ain, we performed transcomplementation experiments with the cloned CS3 1A major-subunit structural genes or cloned cf29A gene and the clp acc essory genes of the CS31A operon. We shelved that CS31A-L, CF29K, and CS31A-K were invoiced in adhesion to Caco-2 and Int-407 cells but not to HEp-2 cells. Nevertheless, K. pneumoniae strains and corresponding E. coli transconjugants producing CF29K adhered to cultured Caco-2, In t-407, and HEp-2 cells, indicating the expression of another R-plasmid -encoded adhesin that mediated adhesion to HEp-2 cells. The carbohydra te part of the eucaryotic receptor of CF29k and CS31A-H adhesins was i nvestigated by adhesion inhibition experiments with Int-407 cells. Alt hough CS31A and CF29K belong to the K88 adhesin family, the receptor d oes not contain N-acetyl-D-galactosamine residues but contains N-acety lneuraminic acid and N-acetyl-D-glucosamine.