Characterization of antithrombins produced by active site mutagenesis of human alpha 1-antitrypsin expressed in yeast

Blood. 1989 Feb;73(2):490-6.

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 alpha 1-antitrypsin (alpha 1AT) cDNA in yeast. Alteration of the alpha 1AT 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 (t1/2) than human alpha 1AT. These antithrombins should be suitable for therapeutic replacement of AT-III in cases of congenital deficiency and in conditions associated with acquired AT-III deficiency, such as disseminated intravascular coagulation.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Antithrombins / genetics*
  • Antithrombins / isolation & purification
  • Antithrombins / metabolism
  • Base Sequence
  • Binding Sites
  • Drug Stability
  • Genetic Variation
  • Humans
  • Kinetics
  • Molecular Sequence Data
  • Mutation*
  • Recombinant Proteins / chemical synthesis*
  • Recombinant Proteins / isolation & purification
  • Recombinant Proteins / metabolism
  • Saccharomyces cerevisiae / genetics*
  • Sequence Homology, Nucleic Acid
  • alpha 1-Antitrypsin / genetics*
  • alpha 1-Antitrypsin / isolation & purification
  • alpha 1-Antitrypsin / metabolism

Substances

  • Antithrombins
  • Recombinant Proteins
  • alpha 1-Antitrypsin