Identification by Tn10 transposon mutagenesis of host factors involved in the biosynthesis of K99 fimbriae of Escherichia coli: Effect of LPS core mutations |
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| Authors: | Emil Pilipcinec Tako T. Huisman Peter T.J. Willemsen Ben J. Appelmelk Frits K. de Graaf Bauke Oudega |
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| Affiliation: | Department of Microbiology and Immunology, University of Veterinary Medicine, Komenského 73, 041 81 Kosice, Slovak Republic; Department of Molecular Microbiology, Faculty of Biology, Vrije Universiteit Amsterdam, De Boelelaan 1087, 1081 HV Amsterdam, the Netherlands; Department of Medical Microbiology, Faculty of Medicine, Vrije Universiteit Amsterdam, the Netherlands |
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| Abstract: | Abstract Tn 10 transposon mutagenesis of Escherichia coli producing K99 fimbriae was carried out to identify host factors involved in regulation or biosynthesis of fimbriae. Two chromosomal mutants were obtained that showed a strongly reduced cell surface expression of K99 fimbriae upon colony blotting and ELISA. Analysis by inversed PCR and nucleotide sequencing showed that one mutant (EP14) contained the Tn 10 transposon in rfaQ , affecting theexpression of the rfaQGP gene cluster, whereas the other mutant (EP35) was affected in a, to date, unknown region of the genome. Immunoblotting analysis confirmed a Rd1 type of LPS of mutant strain EP14. These findings for the first time indicated an effect of LPS core biosynthesis on the biogenesis of fimbriae at the cell surface. Preliminary experiments indicated that K99 major subunits, in contrast to K88 subunits, strongly bind LPS molecules. |
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| Keywords: | Escherichia coli Fimbriae biosynthesis Inversed PCR Lipopolysaccharide core K99 fimbriae RfaQ RfaQ RfaP Tn 10 Transposon mutagenesis LPS binding |
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