Characterization of the F-plasmid conjugative transfer gene traU.
We characterized the traU gene of the Escherichia coli K-12 conjugative plasmid F. Plasmids carrying segments of the F transfer operon were tested for their capacity to complement F lac traU526. The protein products of TraU+ clones were identified, and the nucleotide sequence of traU was determined. traU mapped between traW and trbC. It encodes a 330-amino-acid, Mr36,786 polypeptide that is processed. Ethanol caused accumulation of a precursor polypeptide; removal of ethanol permitted processing of the protein to occur. Because F lac traU526 strains appear to be resistant to F-pilus-specific phages, traU has been considered an F-pilus assembly gene. However, electron microscopic analysis indicated that the traU526 amber mutation caused only a 50% reduction in F-piliation. Since F lac traU526 strains also retain considerable transfer proficiency, new traU mutations were constructed by replacing a segment of traU with a kanamycin resistance gene. Introduction of these mutations into a transfer-proficient plasmid caused a drastic reduction in transfer proficiency, but pilus filaments remained visible at approximately 20% of the wild-type frequency. Like traU526 strains, such mutants were unable to plaque F-pilus-specific phages but exhibited a slight sensitivity on spot tests. Complementation with a TraU+ plasmid restored the wild-type transfer and phage sensitivity phenotypes. Thus, an intact traU product appears to be more essential to conjugal DNA transfer than to assembly of pilus filaments.
Moore
D
D
Department of Medical Microbiology and Immunology, Texas A&M University, College Station 77840.
Maneewannakul
K
K
Maneewannakul
S
S
Wu
J H
JH
Ippen-Ihler
K
K
Bradley
D E
DE
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics*; D001483:Base Sequence; D003001:Cloning, Molecular; D003227:Conjugation, Genetic; D004269:DNA, Bacterial / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics*; D029968:Escherichia coli Proteins; D000431:Ethanol / Q000494:pharmacology; D005144:F Factor; D005798:Genes, Bacterial; D005816:Genetic Complementation Test; D005822:Genetic Vectors; D008969:Molecular Sequence Data; D009154:Mutation; D009876:Operon; D014176:Protein Biosynthesis / Q000187:drug effects; D015183:Restriction Mapping
D001426:Bacterial Proteins; D004269:DNA, Bacterial; D029968:Escherichia coli Proteins; C065100:TraU protein, E coli; D000431:Ethanol