The pilE gene product of Pseudomonas aeruginosa, required for pilus biogenesis, shares amino acid sequence identity with the N-termini of type 4 prepilin proteins.
A new locus required for type 4 pilus biogenesis by Pseudomonas aeruginosa has been identified. A pilE mutant, designated MJ-6, was broadly resistant to pili-specific phages and unable to translocate across solid surfaces by the pilus-dependent mechanism of twitching motility (Twt-). Immunoblot analysis demonstrated that MJ-6 was devoid of pili (Pil-) but was unaffected in the production of unassembled pilin pools. Genetic studies aimed at localizing the pilE mutation on the P. aeruginosa PAO chromosome demonstrated a strong co-linkage between MJ-6 phage resistance and the proB marker located at 71 min. Cloning of the pilE gene was facilitated by the isolation and identification of a pro(B+)-containing plasmid from a PAO1 cosmid library. Upon introduction of the PAO1 proB+ cosmid clone into MJ-6, sensitivity to pili-specific phage, twitching motility and pilus production were restored. The nucleotide sequence of a 1 kb EcoRV-ClaI fragment containing the pilE region revealed a single complete open reading frame with characteristic P. aeruginosa codon bias. PilE, a protein with a molecular weight of 15,278, showed significant sequence identity to the pilin precursors of P. aeruginosa and to other type 4 prepilin proteins. The region of highest homology was localized to the N-terminal 40 amino acid residues. The putative PilE N-terminus contained a seven-residue basic leader sequence followed by a consensus cleavage site for prepilin peptidase and a largely hydrophobic region which contained tyrosine residues (Tyr-24 and Tyr-27) previously implicated in maintaining pilin subunit-subunit interactions. The requirement of PilE in pilus biogenesis was confirmed by demonstrating that chromosomal pilE insertion mutants were pilus- and twitching-motility deficient.
Department of Microbiology, Ohio State University, Columbus 43210.
M A
MA
Russell
A
A
Darzins
D000595:Amino Acid Sequence; D001425:Bacterial Outer Membrane Proteins / Q000737:chemistry*; D001426:Bacterial Proteins / Q000737:chemistry* / Q000378:metabolism; D001483:Base Sequence; D002874:Chromosome Mapping; D002876:Chromosomes, Bacterial; D016384:Consensus Sequence; D010450:Endopeptidases; D036781:Fimbriae Proteins; D010861:Fimbriae, Bacterial / Q000378:metabolism*; D005798:Genes, Bacterial; D005816:Genetic Complementation Test; D008969:Molecular Sequence Data; D011498:Protein Precursors / Q000737:chemistry*; D011550:Pseudomonas aeruginosa / Q000235:genetics*; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid
D001425:Bacterial Outer Membrane Proteins; D001426:Bacterial Proteins; D011498:Protein Precursors; C074063:pilE protein, Neisseria gonorrhoeae; D036781:Fimbriae Proteins; D010450:Endopeptidases; C067127:prepilin peptidase protein, Bacteria