Roll D E, Glew R H
J Biol Chem. 1981 Aug 10;256(15):8190-6.
From 330 ml of rat serum, 222 mg of alpha-1-antitrypsin have been isolated and purified with an overall yield of approximately 20%. The preparation was homogeneous by the criteria of sodium dodecyl sulfate-polyacrylamide gel electrophoresis, sedimentation equilibrium centrifugation, and immunoelectrophoresis. Rat alpha-1-antitrypsin exhibited Mr = 47,000 +/- 1,500 by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and 45,000 +/- 1,000 by equilibrium ultracentrifugation; the sedimentation coefficient (s20,w) was 3.29. Rat alpha-1-antitrypsin exhibited a trypsin-combining ratio (moles of trypsin inhibited/mol of alpha-1-antitrypsin) of 0.88. Rat alpha-1-antitrypsin showed significant differences in amino acid composition when compared to human alpha-1-antitrypsin, particularly in lysine, glutamic acid, arginine, methionine, and tyrosine content. Rat alpha-1-antitrypsin contains 14.3 residues/mol of N-acetylglucosamine, 5.0 residues/mol of mannose, 4.2 residues/mol of galactose, and 5.8 residues/mol of sialic acid. Monospecific antibody produced in a rabbit against our purest preparation of rat alpha-1-antitrypsin does not cross-react immunologically against human, calf, fetal calf, mouse, or chicken sera. The availability of a pure preparation of rat alpha-1-antitrypsin as well as the specific alpha-1-antitrypsin antibody will facilitate studies on the biosynthesis and secretion of this important protease inhibitor in an appropriate animal model.
从330毫升大鼠血清中已分离并纯化出222毫克α-1-抗胰蛋白酶，总产率约为20%。根据十二烷基硫酸钠-聚丙烯酰胺凝胶电泳、沉降平衡离心和免疫电泳的标准，该制剂是均一的。通过十二烷基硫酸钠-聚丙烯酰胺凝胶电泳，大鼠α-1-抗胰蛋白酶的相对分子质量为47,000±1,500，通过平衡超速离心法测得为45,000±1,000；沉降系数（s20,w）为3.29。大鼠α-1-抗胰蛋白酶的胰蛋白酶结合比（抑制的胰蛋白酶摩尔数/α-1-抗胰蛋白酶摩尔数）为0.88。与人类α-1-抗胰蛋白酶相比，大鼠α-1-抗胰蛋白酶在氨基酸组成上有显著差异，尤其是在赖氨酸、谷氨酸、精氨酸、甲硫氨酸和酪氨酸含量方面。大鼠α-1-抗胰蛋白酶每摩尔含有14.3个N-乙酰葡糖胺残基、5.0个甘露糖残基、4.2个半乳糖残基和5.8个唾液酸残基。用兔子针对我们最纯的大鼠α-1-抗胰蛋白酶制剂产生的单特异性抗体，在免疫反应中不与人类、小牛、胎牛、小鼠或鸡的血清发生交叉反应。获得大鼠α-1-抗胰蛋白酶的纯制剂以及特异性α-1-抗胰蛋白酶抗体，将有助于在合适的动物模型中研究这种重要蛋白酶抑制剂的生物合成和分泌。
