title: IS195, an insertion sequence-like element associated with protease genes in Porphyromonas gingivalis.
abstract: Porphyromonas gingivalis is recognized as an important etiologic agent in adult and early-onset periodontal disease. Proteases produced by this organism contribute to its virulence in mice. Protease-encoding genes have been shown to contain multiple copies of repeated nucleotide sequences. These conserved sequences have also been found in hemagglutinin genes. In the process of studying the genetic loci containing the conserved repeated sequences, we have characterized a prtP gene homolog from P. gingivalis W83 encoding a cysteine protease with Lys-X specificity. However, this prtP gene was interrupted by an insertion sequence-like element which we designated IS195. Furthermore, IS195 and another element, IS1126, were present downstream of prtP gene homologs (kgp) found in P. gingivalis H66 and 381. IS195, a 1,068-bp insertion sequence-like element, contained 11-bp inverted repeats at its termini and was bordered by 9-bp direct repeats presumed to be a transposition-mediated target site duplication. Its central region contained one large open reading frame encoding a predicted 300-amino-acid protein which appeared to be a transposase. We isolated two naturally occurring variants of P. gingivalis W83, one carrying IS195 within the coding region of the prtP gene and another containing an intact prtP gene. Biochemical characterization revealed a lack of trypsin-like Lys-X specific proteolytic activity in the P. gingivalis W83 variant carrying the disrupted prtP gene. Studies using a mouse model revealed a reduction of virulence resulting from insertion of IS195 into the coding region of the prtP gene. An allelic-exchange mutant defective in the prtP gene also was constructed and tested in vivo. It displayed intermediate virulence compared to that of the wild-type and prtP::IS195 mutant strains. We conclude that the Lys-X cysteine protease contributes to virulence in soft tissue infections.
authors.affiliation: Department of Microbiology and Immunology, Virginia Commonwealth University, Richmond, Virginia 23298-0678, USA.
authors.forename: J P
authors.initials: JP
authors.lastname: Lewis
authors.forename: F L
authors.initials: FL
authors.lastname: Macrina
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D004251:DNA Transposable Elements; D018628:Gene Dosage; D005798:Genes, Bacterial; D051379:Mice; D008969:Molecular Sequence Data; D016966:Porphyromonas gingivalis / Q000201:enzymology / Q000235:genetics* / Q000472:pathogenicity; D012697:Serine Endopeptidases / Q000235:genetics*; D014774:Virulence
chemical_list: D004251:DNA Transposable Elements; D012697:Serine Endopeptidases; C085783:lactocepin