Osteonectin Degrading Activity in Calf Bone Matrix

Research Project

Project/Area Number	07457498
Research Category	Grant-in-Aid for Scientific Research (B)
Allocation Type	Single-year Grants
Section	一般
Research Field	Surgical dentistry
Research Institution	Tsurumi University
Principal Investigator	ISHIBASHI Katsunori School of Dental Medicine, Tsurumi University Professor, 歯学部, 教授 (20013980)
Co-Investigator(Kenkyū-buntansha)	YAMACHIKA Shigeo School of Dental Medicine, Tsurumi University Assistant, 歯学部, 助手 (60182565)
Project Period (FY)	1995 – 1996
Project Status	Completed (Fiscal Year 1996)
Budget Amount *help	¥6,200,000 (Direct Cost: ¥6,200,000) Fiscal Year 1996: ¥500,000 (Direct Cost: ¥500,000) Fiscal Year 1995: ¥5,700,000 (Direct Cost: ¥5,700,000)
Keywords	calf bone matrix / non-collagenous protein / osteonectin / proteinase
Research Abstract	Soluble proteins were sequentially extracted from demineralized calf bone matrix, with Tris buffer, NaCl solution and guanidine HCl solution. These extracts were concentrated for enzymography, but the guanidine HCl extract was muddied by this manipulation. Then the extract was separated GS (supemate) and GR (residue)-extract by centrifuge. Proteolytic activities in these extracts were investigated by means of enzymograply using gelatin or casein as a substrate. Identical proteolytic active bands were detected on the enzymograms of each the NaCl extract (N-extract), GS-extract and GR-Extract. However, when the purified osteonectin was used as substrate, a proteolytic activity was detected only in the GR-extract. These results would indicate that the presence of a specific osteonectin-lytic proteinase in bone. The enzyme needed Ca ions for activity and was inhibited by EDTA and 1,10-phenanthoroline, it would be an active type metalloproteinase which was bound to bone collagenous matrix Osteonectin degraded fragments were observed 26k, 22k, 20k, 16k, and 14k dalton bands by SDS-PAGE.Sinoe some of these bands were doserved in GR-extract, degraded fragments were bound more tighty than intact osteonectin. This result suggested that osteonectin might be degraded for increase ability of binding to collagen.