Identification by Tn10 transposon mutagenesis of host factors involved in the biosynthesis of K99 fimbriae of Escherichia coli: effect of LPS core mutations.
Tn10 transposon mutagenesis of Escherichia coli producing K99 fimbriae was carried out to identify host factors involved in regulation of 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 Tn10 transposon in rfaQ, affecting the expression 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.
Pilipcinec
E
E
Department of Microbiology and Immunology, University of Veterinary Medicine, Kosice, Slovak Republic.
Huisman
T T
TT
Willemsen
P T
PT
Appelmelk
B J
BJ
de Graaf
F K
FK
Oudega
B
B
D000911:Antibodies, Monoclonal; D001426:Bacterial Proteins / Q000096:biosynthesis / Q000235:genetics*; D001483:Base Sequence; D015342:DNA Probes; D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism; D036781:Fimbriae Proteins; D015964:Gene Expression Regulation, Bacterial; D015151:Immunoblotting; D008070:Lipopolysaccharides / Q000096:biosynthesis* / Q000276:immunology; D008969:Molecular Sequence Data; D016254:Mutagenesis, Insertional; D016133:Polymerase Chain Reaction
D000911:Antibodies, Monoclonal; D001426:Bacterial Proteins; D015342:DNA Probes; D008070:Lipopolysaccharides; C063055:fimbrillin; D036781:Fimbriae Proteins