title: The TadV protein of Actinobacillus actinomycetemcomitans is a novel aspartic acid prepilin peptidase required for maturation of the Flp1 pilin and TadE and TadF pseudopilins.
abstract: The tad locus of Actinobacillus actinomycetemcomitans encodes genes for the biogenesis of Flp pili, which allow the bacterium to adhere tenaciously to surfaces and form strong biofilms. Although tad (tight adherence) loci are widespread among bacterial and archaeal species, very little is known about the functions of the individual components of the Tad secretion apparatus. Here we characterize the mechanism by which the pre-Flp1 prepilin is processed to the mature pilus subunit. We demonstrate that the tadV gene encodes a prepilin peptidase that is both necessary and sufficient for proteolytic maturation of Flp1. TadV was also found to be required for maturation of the TadE and TadF pilin-like proteins, which we term pseudopilins. Using site-directed mutagenesis, we show that processing of pre-Flp1, pre-TadE, and pre-TadF is required for biofilm formation. Mutation of a highly conserved glutamic acid residue at position +5 of Flp1, relative to the cleavage site, resulted in a processed pilin that was blocked in assembly. In contrast, identical mutations in TadE or TadF had no effect on biofilm formation, indicating that the mechanisms by which Flp1 pilin and the pseudopilins function are distinct. We also determined that two conserved aspartic acid residues in TadV are critical for function of the prepilin peptidase. Together, our results indicate that the A. actinomycetemcomitans TadV protein is a member of a novel subclass of nonmethylating aspartic acid prepilin peptidases.
authors.affiliation: Department of Microbiology, College of Physicians and Surgeons, Columbia University, 1516 HHSC, 701 West 168th Street, New York, NY 10032, USA.
authors.forename: Mladen
authors.initials: M
authors.lastname: Tomich
authors.forename: Daniel H
authors.initials: DH
authors.lastname: Fine
authors.forename: David H
authors.initials: DH
authors.lastname: Figurski
mesh_terms: D016976:Aggregatibacter actinomycetemcomitans / Q000201:enzymology* / Q000235:genetics; D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D018441:Biofilms / Q000254:growth & development; D017124:Conserved Sequence; D004252:DNA Mutational Analysis; D010450:Endopeptidases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D036781:Fimbriae Proteins / Q000378:metabolism*; D008969:Molecular Sequence Data; D005810:Multigene Family; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D009876:Operon; D011485:Protein Binding; D011499:Protein Processing, Post-Translational
chemical_list: D001426:Bacterial Proteins; D036781:Fimbriae Proteins; D010450:Endopeptidases; C067127:prepilin peptidase protein, Bacteria