Chemical synthesis, molecular cloning and expression of gene coding for a Bowman-Birk-type proteinase inhibitor

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Abstract
A gene coding for a Bowman-Birk type proteinase inhibitor was synthesized chemically, cloned and expressed in Escherichia coli as a fusion protein with a .beta.-galactosidase fragment. The corresponding mutant inhibitor, carrying a P1 = Arg16 instead of Lys and Ile27 instead of Met was obtained after cyanogen bromide cleavage, refolding and affinity chromatography on trypsin-Sepharose. Dissociation constants of complexes with trypsin of this mutant and wild-type Bowman-Birk inhibitor are identical within experimental error. This is explained by differential patterns of hydrogen bonds between side-chains of Arg or Lys in proteinase inhibitors and the primary specificity pocket of trypsin.