The Amino Acid Sequence of Barley Rootlet Trypsin Inhibitor

Abstract

The amino acid sequence of barley rootlet trypsin inhibitor (BRTI) has been established. The reduced and S-carboxymethylated BRTI was cleaved with cyanogen bromide (CNBr), and three fragments (I, II, and III) were separated using gel filtration on a Bio-Gel P-30 column. Each CNBr-fragment was digested with trypsin and the resulting tryptic peptides were sequenced either by manual Edman degradation or by the DABITC/PITC double-coupling method. Alignment of the tryptic peptides from CNBr-fragment CB-I, which is the largest fragment, was done by analyzing the chymotrypic and thermolytic peptides derived from the CNBr-fragment CB-I. Thus, the amino acid sequence of BRTI consisting of 124 amino acid residues was established. BRTI proved to have intramolecular homology of 55% between the N-terminal half (positions 1 ~62) and the C-terminal half (positions 63 ~ 124); it also displays intermolecular homology with the trypsin inhibitors from wheat germ, rice bran, and soybeans. By comparing with soybean Bowman-Birk trypsin inhibitor, the reactive sites of BRTI are assumed to be Arg(17)-Ser(18) and Arg(75)-Ser(76).