Identification of the noncollagenous proteins of bovine bone by two-dimensional gel electrophoresis.

We have characterized the noncollagenous proteins of bovine bone using two high-resolution gel electrophoretic techniques. Proteins were extracted from bone tissue by extended dialysis against 0.5 M EDTA. In some cases, a preextraction was done in guanidine HCl. Bovine plasma was also examined to identify the proteins in bone that might also be present in blood. We have scored 160 major, reproducible spots on our standard preparation bone map. These comprise about 40 individual protein groups. There are many more minor spots present which puts the total number present over 200. Of these groups, 15 are not present on plasma maps. Bone proteins identified in this way include actin, bone Gla-protein, and osteonectin. The remainder are unknown. Bone Gla-protein is present in bovine bone in four isoelectric forms, pI = 3.95-4.50. Electroblotting analysis of EDTA and guanidine HCl extracted material failed to reveal any higher molecular weight immunologically reactive species. The plasma proteins found in bone include, but are not limited to albumin, apo A-I lipoprotein, IgG, IgM, transferrin, alpha-2-HS-glycoprotein, and hemoglobin. Extraction with guanidine HCl plus EDTA significantly enriches the yield for the nonplasma proteins but does not appear to extract any additional bone proteins.