Studies on Soybean Trypsin Inhibitors

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Abstract
All the known reactive site P1′ positions (notation of Schechter and Berger, 1967) of legume double-headed proteinase inhibitors are occupied by serine residues. To investigate the role of this serine residue, P1′ serine (residue 44) at the chymotrypsin-reactive site of Bowman-Birk inhibitor was replaced with other amino acids by the method of Kowalski and Laskowski, Jr. (1976), using limited proteolysis with chymotrypsin, removal of the serine residue by the Edman degradation method and condensation with activated amino acid esters. By this method six inhibitor derivatives having a Ser, Ala, Thr, Val, Leu, or Gly residue at position 44 were prepared. α-Chymotrypsin inhibitory activity was retained in this order, i.e. the Ser44 derivative was the most active and the Gly44 derivative was essentially inactive to α-chymotrypsin. Gly44-inhibitor prepared by periodate oxidation of Ser44 followed by transamination gave the same result. These results suggest that at least an α-methyl or methylene (methine) group in the P1′ amino acid residue is indispensable for the chymotrypsin inhibitory activity. Bulky side chains in this position appeared to decrease the activity, although a hydroxymethyl side chain was most effective. It is evident that P1′ serine requires conservation for the effective manifestation of the inhibitory activity of the legume double-headed inhibitors.