Separation of bone matrix proteins by calcium-induced precipitation

Calcified Tissue International
Y KubokiK Taniguchi

Abstract

It was found that significant precipitation occurred immediately after calcium, at a concentration as low as 2 mM, was added to a desalted solution of EDTA extract of adult bovine femur. The maximal yield of the precipitates was observed at a calcium concentration of 30 mM. These precipitates were dissolved in 0.5 M EDTA, desalted, and characterized by Sepharose CL-6B gel filtration chromatography and high performance gel-exclusion chromatography. Results revealed that the precipitates were enriched in a 40 K protein and a higher molecular weight fraction as compared with the original extract of bone proteins. The 40 K fraction was isolated and identified as osteonectin, as judged from amino acid analysis, electrophoresis, and immunodetection. The supernatant after calcium-induced precipitation predominantly contained osteocalcin and a 50 K protein that was tentatively identified as alpha 2HS protein. Osteonectin was purified from the calcium-induced precipitates from the EDTA extract of bovine bone. By calcium titration using fluorescence spectrometry, the isolated osteonectin showed high affinity to calcium ions with an apparent dissociation constant (K0.5) of 8 x 10(-7) M. Thus, the use of calcium to separate bone proteins, ...Continue Reading

References

Oct 1, 1975·Proceedings of the National Academy of Sciences of the United States of America·P V HauschkaP M Gallop
May 1, 1976·Proceedings of the National Academy of Sciences of the United States of America·P A PriceN Raman
Jan 1, 1985·Connective Tissue Research·S SatoW T Butler
Jun 15, 1974·European Journal of Biochemistry·B A AshtonG M Herring
Jan 1, 1984·Calcified Tissue International·Y KubokiS Sasaki
Nov 1, 1983·Collagen and Related Research·D Cocking-JohnsonJ J Sauk

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Citations

Jun 1, 1995·Calcified Tissue International·H Takita, Y Kuboki
Aug 14, 1992·Biochemical and Biophysical Research Communications·H Y ZhouY Kuboki

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