Genetic Characterization of the Klebsiella pneumoniae waa Gene Cluster, Involved in Core Lipopolysaccharide Biosynthesis

doi:10.1128/JB.183.12.3564-3573.2001

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Journal of Bacteriology, June 2001, p. 3564-3573, Vol. 183, No. 12
0021-9193/01/$04.00+0 DOI: 10.1128/JB.183.12.3564-3573.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.

Genetic Characterization of the Klebsiella pneumoniae waa Gene Cluster, Involved in Core Lipopolysaccharide Biosynthesis

Miguel Regué,¹^,^* Núria Climent,¹ Nihal Abitiu,¹ Núria Coderch,¹ Susana Merino,² Luis Izquierdo,² Maria Altarriba,² and Juan M. Tomás²

Departamento de Microbiología y Parasitología Sanitarias, División de Ciéncias de la Salud, Facultad de Farmacia,¹ and Departamento de Microbiología, Facultad de Biología,² Universidad de Barcelona, Barcelona, Spain

Received 22 January 2001/Accepted 4 April 2001

A recombinant cosmid containing genes involved in Klebsiella pneumoniae C3 core lipopolysaccharide biosynthesis was identifiedby its ability to confer bacteriocin 28b resistance to Escherichiacoli K-12. The recombinant cosmid contains 12 genes, the wholewaa gene cluster, flanked by kbl and coaD genes, as was foundin E. coli K-12. PCR amplification analysis showed that this clusteris conserved in representative K. pneumoniae strains. Partialnucleotide sequence determination showed that the same genes andgene order are found in K. pneumoniae subsp. ozaenae, for whichthe core chemical structure is known. Complementation analysisof known waa mutants from E. coli K-12 and/or Salmonella entericaled to the identification of genes involved in biosynthesis ofthe inner core backbone that are shared by these three membersof the Enterobacteriaceae. K. pneumoniae orf10 mutants showeda two-log-fold reduction in a mice virulence assay and a strongdecrease in capsule amount. Analysis of a constructed K. pneumoniaewaaE deletion mutant suggests that the WaaE protein is involvedin the transfer of the branch $beta$ -D-Glc to the O-4 position of L-glycero-D-manno-heptoseI, a feature shared by K. pneumoniae, Proteus mirabilis, and Yersiniaenterocolitica.

^* Corresponding author. Mailing address: Departamento de Microbiología y Parasitología Sanitarias, Facultad de Farmacia, Universidadde Barcelona, Av. Joan XXIII s/n, Barcelona 08028, Spain. Phone:34-3-4024496. Fax: 34-3-4021886. E-mail: regue{at}farmacia.far.ub.es.

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