Characterization of antithrombins produced by active site mutagenesis of human a1-antitrypsin expressed in yeast

Abstract

Both congenital and acquired antithrombin-III (AT-III) deficiencies are amenable to replacement therapy. We describe two antithrombins produced by recombinant DNA techniques from human a1-antitrypsin (a1AT) cDNA in yeast. Alteration of the a1AT active site, replacing methionine 358 with arginine, results in a thrombin inhibition rate similar to that of heparin-activated AT-III. Alteration of two further residues, to give a five-residue sequence identical to AT-III, does not increase this rate further. Neither antithrombin is activated by heparin; both are unglycosylated and have shorter in vivo half-lives (t 1/2 ) than human a1AT. These antithrombins should be suitable for therapeutic replacement of AT-III in cases of conginetal deficiency and in conditions associated with acquired AT-III deficiency, such as disseminated intravascular coagulation.