Purification of a proteinase inhibitor from bovine serum with C1-inhibitor activity.
This report describes the purification of a novel proteinase inhibitor from bovine serum. This protein was purified to apparent homogeneity employing affinity binding to sulfated dextran and precipitation by ammonium sulfate, followed by sequential chromatography on DEAE-cellulose, heparin-Sepharose and Sephacryl S-200. Quantitative enzyme-linked immunosorbent assays revealed that the concentration of this inhibitor is approximately 3 microM in bovine serum. The inhibitor is a single polypeptide chain with an estimated Mr of 83,000 as determined by SDS-polyacrylamide gel electrophoresis. An aspartic acid was found at the amino terminus of the protein; N-terminal amino acid sequence data indicated that there was no significant homology with other reported amino acid sequences. This bovine inhibitor covalently complexed the human proteinases C1-r, C1-s, factor XIIa and plasma kallikrein, which are also complexed and inactivated by human C1-inhibitor. In addition, the bovine inhibitor complexed and inactivated bovine chymotrypsin, a feature which functionally distinguishes it from human C1-inhibitor. Although the bovine inhibitor appears functionally very similar to C1-inhibitor, we found no evidence for structural homology with the human counterpart.
D000595:Amino Acid Sequence; D000818:Animals; D002417:Cattle; D003174:Complement C1 Inactivator Proteins / Q000097:blood* / Q000302:isolation & purification / Q000502:physiology; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D010449:Peptide Mapping; D011480:Protease Inhibitors / Q000097:blood* / Q000302:isolation & purification / Q000502:physiology; D013379:Substrate Specificity
D003174:Complement C1 Inactivator Proteins; D011480:Protease Inhibitors