Isolation and characterization of the Porphyromonas gingivalis prtT gene, coding for protease activity.
The prtT gene, coding for trypsinlike proteolytic activity, has been isolated from Porphyromonas gingivalis ATCC 53977. This gene is present immediately downstream from the sod gene on a 5.9-kb DNA fragment from the organism isolated in Escherichia coli. The complete nucleotide sequence of the gene was determined, and the deduced amino acid sequence of the enzyme corresponds to a 53.9-kDa protein with an estimated pI of 11.85. Gelatin-sodium dodecyl sulfate-polyacrylamide gel electrophoresis zymography also indicated a similar molecular size for the protease. The enzyme was purified to near homogeneity following anion-exchange and gel-filtration chromatography. The purified enzyme also exhibited a single protein species with a size of approximately 53 kDa. Enzyme activity was strongly dependent upon the presence of reducing agents (dithiothreitol, cysteine, and 2-mercaptoethanol) and was also stimulated in the presence of calcium ions. A comparison of the properties of the prtT gene product with comparable parameters of proteases previously purified from different strains of P. gingivalis suggested that the cloned protease represents a previously uncharacterized enzyme.
Otogoto
J
J
Department of Pediatric Dentistry, University of Texas Health Science Center, San Antonio 78284-7888.
Kuramitsu
H K
HK
D000595:Amino Acid Sequence; D001426:Bacterial Proteins; D001483:Base Sequence; D003001:Cloning, Molecular; D003546:Cysteine Endopeptidases; D004591:Electrophoresis, Polyacrylamide Gel; D005798:Genes, Bacterial; D008969:Molecular Sequence Data; D012150:Polymorphism, Restriction Fragment Length; D016966:Porphyromonas gingivalis / Q000235:genetics*; D017422:Sequence Analysis, DNA; D014357:Trypsin / Q000235:genetics* / Q000302:isolation & purification
D001426:Bacterial Proteins; D014357:Trypsin; D003546:Cysteine Endopeptidases; C078912:prtT protein, Porphyromonas gingivalis