Cloning, expression, purification, and biological activity of recombinant native and variant human alpha 1-antichymotrypsins

J Biol Chem. 1990 Jan 15;265(2):1199-207.


Human alpha 1-antichymotrypsin has been cloned, sequenced and expressed in Escherichia coli and recombinant protein as well as point-specific mutants have been purified and characterized. The corrected gene-deduced amino acid sequence has 45% overall identity with alpha 1-protease inhibitor, which is higher than the 42% previously reported (Chandra, T., Stackhouse, R., Kidd, V. J., Robson, J. H., and Woo, S. L. C. (1983) Biochemistry 22, 5055-5060). Recombinant antichymotrypsin (rACT) is similar to natural antichymotrypsin with respect to the specificity of its interactions with proteases. Its second-order rate constant for association with bovine chymotrypsin is 6-8 x 10(5) M-1 s-1, which is identical to that of the serum-derived inhibitor. Site-specific mutagenesis has been used to produce two variants of rACT in which the P1 position has been changed from leucine to either methionine (L358M-rACT) or arginine (L358R-rACT). L358M-rACT has a specificity of inhibitory activity toward serine proteases closely similar to that of native rACT. By contrast, the specificity of L358R-rACT is quite different from that of native rACT, most notably in efficiently inhibiting trypsin and human thrombin while showing a decreased ability to inhibit chymotrypsin.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Base Sequence
  • Blotting, Western
  • Cloning, Molecular*
  • Electrophoresis, Polyacrylamide Gel
  • Escherichia coli / genetics
  • Gene Expression*
  • Genes, Bacterial
  • Humans
  • Kinetics
  • Molecular Sequence Data
  • Mutation
  • Recombinant Proteins / genetics
  • Swine
  • alpha 1-Antichymotrypsin / genetics*
  • alpha 1-Antichymotrypsin / isolation & purification
  • alpha 1-Antichymotrypsin / metabolism


  • Recombinant Proteins
  • alpha 1-Antichymotrypsin

Associated data

  • GENBANK/J05176