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. 1993 Oct;61(10):4139–4146. doi: 10.1128/iai.61.10.4139-4146.1993

Characterization of the tpr gene product and isolation of a specific protease-deficient mutant of Porphyromonas gingivalis W83.

Y Park 1, B C McBride 1
PMCID: PMC281136  PMID: 8406803

Abstract

The previously described protease gene (tpr) of Porphyromonas gingivalis W83 was shown by sodium dodecyl sulfate-polyacrylamide gel electrophoresis of the recombinant protein and in vitro translation to encode a 50-kDa protein whose active form migrates with an apparent molecular mass of 90 kDa. The 50-kDa protein was expressed at high levels by using a T7 RNA polymerase/promoter system. The NH2-terminal sequence of the protein was identical to the amino acid sequence deduced from the DNA sequence of the protease gene. Affinity-purified antibody to the 90-kDa recombinant protease reacted with an 80-kDa P. gingivalis protein. A specific protease (Tpr)-deficient isogenic mutant of P. gingivalis was generated by homologous recombination between P. gingivalis chromosomal DNA and a suicide plasmid carrying the cloned gene disrupted by insertion of an erythromycin resistance gene. Gelatin substrate zymography showed that cell extracts of the mutant lacked a protease band that migrated with an apparent molecular mass of 80 kDa. Western immunoblots of the cell extracts indicated the loss of an antigen with a similar mass.

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Selected References

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