Role of bacterial γ-glutamyltranspeptidase as a novel virulence factor in bone-resorbing pathogenesis.
Mammalian γ-glutamyltranspeptidase (GGT) has been identified as a bone-resorbing factor. Since GGT of Bacillus subtilis exhibits similarity in their primary structure and enzymatic characteristics with mammalian GGTs, the bone-resorbing activity of bacterial GGT was examined in this study. Osteoclastogenesis was performed in a co-culture system of mouse calvaria-derived osteoblasts and bone marrow cells. A conditioned medium from GGT-overproducing B. subtilis culture showed significantly higher activity of osteoclast formation than a conditioned medium from wild-type B. subtilis culture. Recombinant GGT (rGGT) of wild-type B. subtilis and an enzymatic activity-defected rGGT of B. subtilis 2288 mutant were expressed in Escherichia coli and purified using His tag. Both purified rGGTs induced similar levels of osteoclastogenesis, suggesting that B. subtilis GGT possesses virulent bone-resorbing activity and its activity is probably independent of its enzymatic activity. Furthermore, a recombinant protein of B. subtilis GGT heavy subunit (Bs rGGT/H) showed strong activity of osteoclastogenesis while the light subunit failed to show strong activity, suggesting that the bone-resorbing activity is mainly located at the heavy subunit. More importantly, the GGT enzymatic activity may not be required for this virulence activity since the light subunit contains the catalytic pocket. In addition, B. subtilis rGGT stimulated mRNA expressions of receptor activator of nuclear factor kappa-B ligand (RANKL) and cyclooxygenase-2 (COX-2), while an osteoprotegerin inhibited the osteoclast formation induced by Bs rGGT/H. This is the first demonstration that bacterial GGT itself is sufficient to act as a bone-resorbing virulence factor via RANKL-dependent pathway. Therefore, it can be hypothesized that GGT of periodontopathic bacteria may play an important role as a virulence factor in bone destruction.
Department of Oral Biology, Oral Science Research Center, BK21 Plus Project, Yonsei University College of Dentistry, Seoul, 03722, Republic of Korea.
Jinmoon
J
Kim
Department of Oral Biology, Oral Science Research Center, BK21 Plus Project, Yonsei University College of Dentistry, Seoul, 03722, Republic of Korea.|Department of Applied Life Science, The Graduate School, Yonsei University, Seoul, 03722, Republic of Korea.
Sungil
S
Jang
Department of Oral Biology, Oral Science Research Center, BK21 Plus Project, Yonsei University College of Dentistry, Seoul, 03722, Republic of Korea.|Department of Applied Life Science, The Graduate School, Yonsei University, Seoul, 03722, Republic of Korea.
Aeryun
A
Kim
Department of Oral Biology, Oral Science Research Center, BK21 Plus Project, Yonsei University College of Dentistry, Seoul, 03722, Republic of Korea.|Department of Applied Life Science, The Graduate School, Yonsei University, Seoul, 03722, Republic of Korea.
Hanfu
H
Su
Department of Oral Biology, Oral Science Research Center, BK21 Plus Project, Yonsei University College of Dentistry, Seoul, 03722, Republic of Korea.
Niluka
N
Gunawardhana
Department of Oral Biology, Oral Science Research Center, BK21 Plus Project, Yonsei University College of Dentistry, Seoul, 03722, Republic of Korea.
Yeong-Eui
YE
Jeon
Department of Oral Biology, Oral Science Research Center, BK21 Plus Project, Yonsei University College of Dentistry, Seoul, 03722, Republic of Korea.
Eun Jung
EJ
Bak
Department of Dental Hygiene, Jeonju Kijeon College, Jeonju, 560-701, Republic of Korea. hippo6@kijeon.ac.kr.
Ji-Hye
JH
Kim
Department of Oral Biology, Oral Science Research Center, BK21 Plus Project, Yonsei University College of Dentistry, Seoul, 03722, Republic of Korea. Jcha@yuhs.ac.|Department of Applied Life Science, The Graduate School, Yonsei University, Seoul, 03722, Republic of Korea. Jcha@yuhs.ac.
Jeong-Heon
JH
Cha
D000818:Animals; D001412:Bacillus subtilis / Q000201:enzymology*; D001854:Bone Marrow Cells / Q000187:drug effects; D001862:Bone Resorption / Q000139:chemically induced / Q000382:microbiology / Q000473:pathology; D018920:Coculture Techniques; D016207:Cytokines / Q000378:metabolism; D051379:Mice; D010006:Osteoblasts / Q000187:drug effects; D010010:Osteoclasts / Q000187:drug effects; D010012:Osteogenesis / Q000187:drug effects*; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology; D037521:Virulence Factors / Q000235:genetics / Q000494:pharmacology / Q000502:physiology; D005723:gamma-Glutamyltransferase / Q000235:genetics / Q000494:pharmacology* / Q000502:physiology
D016207:Cytokines; D011994:Recombinant Proteins; D037521:Virulence Factors; C025940:bone resorption factor; D005723:gamma-Glutamyltransferase
Bacillus subtilis; bone resorption; osteoclastogenesis; γ-glutamyltranspeptidase