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Putative down-stream signaling molecule of GTPase in Porphyromonas gingivalis

Putative down-stream signaling molecule of GTPase in Porphyromonas gingivalis Porphyromonas gingivalis is a strict anaerobic bacterium mainly responsible for periodontal disease in oral cavity. Putative GTPase gene (pgp) of this bacterium was cloned and its recombinant protein (rPGP) was produced in Escherichia coli. Based on the amino acid sequence of SGP that is a GTP-binding protein of Streptococcus mutans, putative GTPase amino acid sequence was deduced in the data base of genome sequences of Porphyromonas gingivalis. A 900-bp PCR fragment was amplified with P. gingivalis genomic DNA as a template and cloned into E. coli JM109. Then pgp was transferred into pQE-30 expression vector to make pQE-PGP for production of rPGP. This protein was produced and purified by Ni-NTA affinity column chromatography. Anti-PGP antibody was also produced in Sprague Dawley rats. Using Westernblot analysis with this antibody, it was confirmed that the rPGP produced in E. coli was identical to that of donor strain. Furthermore, by Southernblot analysis it was revealed that the pgp was originated from P. gingivalis. By immunoprecipitation with anti-PGP antibody and N-terminal amino acid sequence analysis it was found that PGP was able to bind to acetate kinase, which was reported to be a secondary signaling molecule in anaerobic microorganisms. Therefore, these results imply that P. gingivalis produces putative GTPase and this protein might play a potential role in signaling pathway in oral biofilm formation. http://www.deepdyve.com/assets/images/DeepDyve-Logo-lg.png Applied Biochemistry and Microbiology Springer Journals

Putative down-stream signaling molecule of GTPase in Porphyromonas gingivalis

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References (1)

Publisher
Springer Journals
Copyright
Copyright © 2012 by Pleiades Publishing, Ltd.
Subject
Life Sciences; Medical Microbiology; Biochemistry, general; Microbiology
ISSN
0003-6838
eISSN
1608-3024
DOI
10.1134/S0003683812030039
Publisher site
See Article on Publisher Site

Abstract

Porphyromonas gingivalis is a strict anaerobic bacterium mainly responsible for periodontal disease in oral cavity. Putative GTPase gene (pgp) of this bacterium was cloned and its recombinant protein (rPGP) was produced in Escherichia coli. Based on the amino acid sequence of SGP that is a GTP-binding protein of Streptococcus mutans, putative GTPase amino acid sequence was deduced in the data base of genome sequences of Porphyromonas gingivalis. A 900-bp PCR fragment was amplified with P. gingivalis genomic DNA as a template and cloned into E. coli JM109. Then pgp was transferred into pQE-30 expression vector to make pQE-PGP for production of rPGP. This protein was produced and purified by Ni-NTA affinity column chromatography. Anti-PGP antibody was also produced in Sprague Dawley rats. Using Westernblot analysis with this antibody, it was confirmed that the rPGP produced in E. coli was identical to that of donor strain. Furthermore, by Southernblot analysis it was revealed that the pgp was originated from P. gingivalis. By immunoprecipitation with anti-PGP antibody and N-terminal amino acid sequence analysis it was found that PGP was able to bind to acetate kinase, which was reported to be a secondary signaling molecule in anaerobic microorganisms. Therefore, these results imply that P. gingivalis produces putative GTPase and this protein might play a potential role in signaling pathway in oral biofilm formation.

Journal

Applied Biochemistry and Microbiology – Springer Journals

Published: Jul 4, 2012

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