Cloning and characterization of a new protease gene (prtH) from Porphyromonas gingivalis.
Porphyromonas gingivalis has been implicated as a contributing etiological agent of adult periodontitis and generalized forms of early-onset periodontitis. Proteases of P. gingivalis may contribute to its pathogenicity by destroying connective tissue as well as inactivating key plasma proteins that might mediate protective host functions. In order to explore this problem, antiserum raised against membrane vesicles of P. gingivalis W83 was used to screen a genomic library of strain W83 constructed by using the lambda DASH vector system. A recombinant phage (lambda 34) expressing a P. gingivalis protease from the library was identified and characterized. Casein substrate zymography of lambda 34 lysates revealed a protease with an apparent molecular mass of 97 kDa. The gene encoding this protease was designated prtH. It was localized to a 3.7-kb HindIII-BamHI fragment and specified an enzyme which hydrolyzed the human C3 complement protein under defined conditions. The nucleotide sequence of this 3.7-kb fragment was determined, and one 2.9-kb open reading frame (992 amino acids) corresponding to a 110-kDa protein was detected, suggesting it might be a precursor of the 97-kDa active protease. prtH is not similar to any previously cloned protease gene from P. gingivalis.
Fletcher
H M
HM
Department of Microbiology and Immunology, Virginia Commonwealth University, Richmond 23298-0678.
Schenkein
H A
HA
Macrina
F L
FL
D000595:Amino Acid Sequence; D000818:Animals; D001426:Bacterial Proteins; D001483:Base Sequence; D002364:Caseins / Q000378:metabolism; D003546:Cysteine Endopeptidases / Q000737:chemistry / Q000235:genetics*; D010450:Endopeptidases / Q000032:analysis / Q000737:chemistry / Q000235:genetics*; D005798:Genes, Bacterial; D008969:Molecular Sequence Data; D010957:Plasmids; D016966:Porphyromonas gingivalis / Q000201:enzymology / Q000235:genetics*; D011817:Rabbits; D011994:Recombinant Proteins / Q000032:analysis
D001426:Bacterial Proteins; D002364:Caseins; D011994:Recombinant Proteins; D010450:Endopeptidases; D003546:Cysteine Endopeptidases; C089744:PrtH protein, bacteria