The isolation of alpha-1-protease inhibitor by a unique procedure designed for industrial application

Abstract

Individuals who are congenitally deficient in the human plasma protein α 1-protease inhibitor ( α 1PI, which is also called α 1-antitrypsin) usually develop chronic obstructive lung disease as a consequence of improperly regulated granulocyte elastase. In this report, a unique, facile one- or two-step method is presented for the large-scale isolation of α 1PI for potential therapeutic use. The method takes advantage of the unusual disulfide bond in α 1PI, which consists of a single cysteine residue in the polypeptide chain bound to a free pendant cysteine. In contrast to other circulating plasma proteins, the disulfide bridge in α 1PI does not add to its structural stability. Therefore, if an α 1PI-containing solution of plasma proteins is precipitated out in the presence of reductant, much more extensive separation of contaminating proteins will be achieved than in the absence of such reductant. We have used Cohn Fraction IV-1, a relatively unused side product in albumin and gamma globulin production, as our starting material. After activation of the α 1PI in basic media, partial purification is achieved with successive additions of Aerosil (a fumed silica), dithiothreitol, and ammonium sulfate. From 90 to 95% of the contaminating proteins are precipitated by this single procedure, resulting in a product that is ∼70% pure. DEAE-cellulose chromatography can be used as an additional purification step, and this results in a product that is nearly homogenous. Overall yield is ∼45%. The method is simple, inexpensive, and reproducible and is directly applicable to large-scale industrial processing.