TY - JOUR
T1 - Bovine Dentin Phosphophoryn
T2 - Composition and Molecular Weight
AU - Stetler-Stevenson, W. G.
AU - Veis, Arthur
PY - 1983/1/1
Y1 - 1983/1/1
N2 - The molecular weight of phosphophoryn, an acidic phosphoprotein unique to dentin matrix, has been difficult to determine because of a combination of neutral protease activities in this tissue and the intrinsic high charge density of the molecule. In this study, bovine dentin phosphophoryn (BDPP) was isolated by a procedure designed to prevent proteolysis. Bovine unerupted third molar powder was demineralized by ethylenediaminetetraacetic acid (EDTA). The EDTA-soluble phosphophoryn fraction was isolated and purified by sequential calcium chloride precipitation, gel filtration in sodium dodecyl sulfate (NaDodSO4) containing buffer, anion-exchange chromatography, and finally gel filtration in 4 M guanidine hydrochloride (4 M Gdn·HCl) buffer. Sedimentation equilibrium, sedimentation velocity, and diffusion coefficient data, viscosity studies in a high ionic strength buffer, and NaDodSO4 gradient gel electrophoresis data gave consistent results for the molecular weight of BDPP, all being in the range of 151 000–167 000. This range is much higher than any previously reported value. An anomalous behavior was observed in nongradient NaDodSO4 gel electrophoresis. Dissociative analytical gel filtration chromatography in 4 M Gdn·HCl gave a molecular weight value of 100 000. This discrepancy was resolved by studying the viscosity of BDPP in 4 M Gdn·HCl which showed BDPP does not assume a true random-chain conformation in this solvent.
AB - The molecular weight of phosphophoryn, an acidic phosphoprotein unique to dentin matrix, has been difficult to determine because of a combination of neutral protease activities in this tissue and the intrinsic high charge density of the molecule. In this study, bovine dentin phosphophoryn (BDPP) was isolated by a procedure designed to prevent proteolysis. Bovine unerupted third molar powder was demineralized by ethylenediaminetetraacetic acid (EDTA). The EDTA-soluble phosphophoryn fraction was isolated and purified by sequential calcium chloride precipitation, gel filtration in sodium dodecyl sulfate (NaDodSO4) containing buffer, anion-exchange chromatography, and finally gel filtration in 4 M guanidine hydrochloride (4 M Gdn·HCl) buffer. Sedimentation equilibrium, sedimentation velocity, and diffusion coefficient data, viscosity studies in a high ionic strength buffer, and NaDodSO4 gradient gel electrophoresis data gave consistent results for the molecular weight of BDPP, all being in the range of 151 000–167 000. This range is much higher than any previously reported value. An anomalous behavior was observed in nongradient NaDodSO4 gel electrophoresis. Dissociative analytical gel filtration chromatography in 4 M Gdn·HCl gave a molecular weight value of 100 000. This discrepancy was resolved by studying the viscosity of BDPP in 4 M Gdn·HCl which showed BDPP does not assume a true random-chain conformation in this solvent.
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U2 - 10.1021/bi00287a025
DO - 10.1021/bi00287a025
M3 - Article
C2 - 6414510
AN - SCOPUS:0021115826
SN - 0006-2960
VL - 22
SP - 4326
EP - 4335
JO - Biochemistry
JF - Biochemistry
IS - 18
ER -