Comparative Studies of High MrSubunits of Rye and Wheat. II. Partial Amino Acid Sequences

Abstract

High M. subunit fractions of two rye cultivars (Danko and Halo) and of the wheat cultivar Rektor were isolated from defatted flours by extraction with 50% (v/v) aqucous propan-1-ol under reducing conditions and 60 °C followed by precipitation using a 60% concentration of propan-1-ol, centrifugation, dialysis and lyophilisation. Two single major subunits of rye Danko were isolated by to wheat high M r subunits. Total high M r subunit fractions of Danko, Halo and Rektor were digested with TPCK-treated trypsin at 37°C for 8h. The partial hydrolysates were pre-separated into the fractions I (M. 10 000) by centrifugation on a Centriprep 10-membranc. Both fractions were then separated by reversed-phase HPLC on C 18 , silica gel. Dominating peptides were purified by preparative rechromatography under slightly changed HPLC conditions and characterised by amino acid analysis and determination of amino acid sequences. The resulting sequences of 19 (Danko), 12 (Halo) and 15 peptides (Rektor) were compared with known sequences from x- and y-type high M r subunits of wheat. With the exception of three peptides, the sequences could be assigned to the three different domains of high M r subunits. Even when the sequences of the rye peptides were similar to those of wheat, they were not identical in any case; on average the variations were about 20%. Most of rye peptides corresponded to the x-type subunits of wheat. Within the most characteristic sequence motifs belonging to the repetitive domain B, fundamental differences between wheat and tye were not detected.