Purification and characterization of rat plasma alpha-1-antitrypsin.

alpha-1-Antitrypsin was purified from rat plasma using affinity chromatography on Cibacron Blue-agarose, ion-exchange chromatography on DE-52 (pH 8.5 and 6.0) and gel filtration on Sephadex G-150. The final preparation was homogeneous as judged by polyacrylamide gel electrophoresis in the presence and absence of sodium dodecyl sulfate, by immunoelectrophoresis and by analytical ultracentrifugation. It was composed of a single polypeptide chain whose molecular weight was estimated to be about 50,000 by sedimentation equilibrium analysis. Rat alpha-1-antitrypsin was shown to be a glycoprotein containing 3.3% glucosamine, 1.7% mannose, 1.0% galactose, and 4.2% sialic acid. The interaction of rat alpha-antitrypsin with bovine beta-trypsin was studied by protease activity assays and by gel electrophoresis in sodium dodecyl sulfate. Rat alpha-1-antitrypsin inhibited bovine beta-trypsin by forming a stable equimolar complex concomitant with the release of a peptide with a molecular weight of approx. 8,000.