Lipopolysaccharide biosynthesis-related genes are required for colony pigmentation of Porphyromonas gingivalis

• Published in: Microbiology (Society for General Microbiology) Vol. 155; no. 4; pp. 1282 - 1293
• Main Authors: Sato, Keiko, Kido, Nobuo, Murakami, Yukitaka, Hoover, Charles I, Nakayama, Koji, Yoshimura, Fuminobu
• Format: Journal Article
• Language: English
• Published: Reading Soc General Microbiol 01.04.2009; Society for General Microbiology
• Subjects: Adhesins, Bacterial - metabolism; Bacterial Proteins - genetics; Bacterial Proteins - metabolism; Bacteriology; Biological and medical sciences; Culture Media; Cysteine Endopeptidases - metabolism; DNA Transposable Elements; Fundamental and applied biological sciences. Psychology; Gene Expression Regulation, Bacterial; Genetics; Gingipain Cysteine Endopeptidases; Glycosyltransferases - genetics; Glycosyltransferases - metabolism; Heme - metabolism; Lipopolysaccharides - biosynthesis; Microbiology; Mutagenesis, Insertional; Pigmentation - genetics; Pigmentation - physiology; Porphyromonas gingivalis; Porphyromonas gingivalis - enzymology; Porphyromonas gingivalis - genetics; Porphyromonas gingivalis - growth & development; Porphyromonas gingivalis - physiology; Uridine Diphosphate Glucose Dehydrogenase - genetics; Uridine Diphosphate Glucose Dehydrogenase - metabolism; Porphyromonas gingivalis; Gene; Lipopolysaccharide; Bacteria; Biosynthesis; Pigmentation; Bacteroidaceae
• ISSN: 1350-0872; 1465-2080
• DOI: 10.1099/mic.0.025163-0

1 Department of Microbiology, School of Dentistry, Aichi-Gakuin University, 1-100 Kusumoto-cho, Chikusa-ku, Nagoya, Aichi 464-8650, Japan 2 Division of Plant Growth Physiology, Nagoya University Graduate School of Biological Sciences, Furou-cho, Chikusa-ku, Nagoya, Aichi 464-8602, Japan 3 University of California, San Francisco, CA 94143-0512, USA 4 Division of Microbiology and Oral Infection, Department of Molecular Microbiology and Immunology, Nagasaki University Graduate School of Biomedical Sciences, 1-7-1 Sakamoto, Nagasaki, Nagasaki 852-8588, Japan Correspondence Keiko Sato satou{at}nagasaki-u.ac.jp The periodontopathic bacterium Porphyromonas gingivalis forms pigmented colonies when incubated on blood agar plates as a result of accumulation of µ-oxo haem dimer on the cell surface. Gingipain–adhesin complexes are responsible for production of µ-oxo haem dimer from haemoglobin. Non-pigmented mutants (Tn6-5, Tn7-1, Tn7-3 and Tn10-4) were isolated from P. gingivalis by Tn 4351 transposon mutagenesis [Hoover & Yoshimura (1994), FEMS Microbiol Lett 124 , 43–48]. In this study, we found that the Tn6-5, Tn7-1 and Tn7-3 mutants carried Tn 4351 DNA in a gene homologous to the ugdA gene encoding UDP-glucose 6-dehydrogenase, a gene encoding a putative group 1 family glycosyltransferase and a gene homologous to the rfa gene encoding ADP heptose-LPS heptosyltransferase, respectively. The Tn10-4 mutant carried Tn 4351 DNA at the same position as that for Tn7-1. Gingipain activities associated with cells of the Tn7-3 mutant ( rfa ) were very weak, whereas gingipain activities were detected in the culture supernatants. Immunoblot and mass spectrometry analyses also revealed that gingipains, including their precursor forms, were present in the culture supernatants. A lipopolysaccharide (LPS) fraction of the rfa deletion mutant did not show the ladder pattern that was usually seen for the LPS of the wild-type P. gingivalis . A recombinant chimera gingipain was able to bind to an LPS fraction of the wild-type P. gingivalis in a dose-dependent manner. These results suggest that the rfa gene product is associated with biosynthesis of LPS and/or cell-surface polysaccharides that can function as an anchorage for gingipain–adhesin complexes. Abbreviations: ABTS, 2,2'-azino-bis(3-ethylbenzthiazoline-6-sulfonic acid) Present address: Division of Microbiology and Oral Infection, Department of Molecular Microbiology and Immunology, Nagasaki University Graduate School of Biomedical Sciences, 1-7-1 Sakamoto, Nagasaki, Nagasaki 852-8588, Japan. Supplementary tables of strains and primers, and details of the construction of the rfa insertion mutant, including a supplementary figure, are available with the online version of this paper.