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Cytotechnology

, Volume 31, Issue 1–2, pp 53–60 | Cite as

A recombinant bait region mutant of human α2-macroglobulin exhibiting an altered proteinase-inhibiting spectrum

  • Atsushi Ikai
  • Kayoko Ookata
  • Masaru Shimizu
  • Noboru Nakamichi
  • Mamiko Ito
  • Toshiharu Matsumura
Article

Abstract

Alpha 2-macroglobulin (α2M), a plasma glycoprotein produced in the liver, inhibits a variety of proteinases and thus considered to play important homeostatic roles in the body. This broad inhibitory spectrum has been explained by the trapping theory by which a proteinase recognizes a region of 25–30 amino acid peptide in α2M called bait region and cleaves it, leading to the conformational change of α2M, and to the subsequent entrapment and inhibition of the proteinase. We constructed α2M cDNAs with mutated DNA sequences in the bait region, and obtained recombinant CHO cell lines producing either wild type α2M, or mutant α2Ms, i.e., α2M/K692 and α2M/K696, each with substitution of Arg with Lys at codons 692 and 696, respectively. We tested if lysyl endopeptidase is not inhibited by wild type α2M, but could be inhibited by these engineered mutant α2Ms. Thus, recombinant α2M/K696 protein successfully inhibited lysyl endopeptidase activity, while recombinant α2M/K692 protein was not sensitive to lysyl endopeptidase, suggesting that not all bait region peptide bonds can equally be accessible and susceptible to proteinases. The present results not only provided the trapping theory with additional supportive evidence, but the first experimental evidence for the value of engineered α2M-derived proteinase inhibitor with an artificial proteinase inhibitory spectrum of potential industrial and/or therapeutic usefulness.

alpha-2-macroglobulin α2M bait region CHO cells human lysyl endopeptidase 

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Copyright information

© Kluwer Academic Publishers 1999

Authors and Affiliations

  • Atsushi Ikai
  • Kayoko Ookata
  • Masaru Shimizu
  • Noboru Nakamichi
  • Mamiko Ito
  • Toshiharu Matsumura

There are no affiliations available

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