CHARACTERIZATION OF THE SUBUNITS OF GLYCININ AND THE COMPLETE PRIMARY STRUCTURE OF THE A2B1A SUBUNIT (SOYBEAN)
Abstract
Five disulfide linked subunits, each consisting of an acidic and a basic polypeptide, were purified from unreduced S-pyridylethylated glycinin from soybean {Glycine max (L.) Merr.} cultivar CX635-1-1-1. Each polypeptide was identified by a combination of NH(,2)-terminal sequence analysis, sodium dodecylsulfate (SDS)-electrophoresis, and analytical isoelectric focusing. Components of each subunit were A(,1a)B(,2), A(,1b)B(,1b), A(,2)B(,1a), A(,3)B(,4) and A(,5)B(,3). A single cystine in analogous positions in each subunit was responsible for linking the acidic and basic chains. A(,4) was not covalently linked to a basic chain. The specific pairing between chains suggested that each glycinin subunit was synthesized as a precursor protein which underwent proteolytic cleavage to yield the disulfide linked acidic and basic components. The complete amino acid sequence was determined for each chain of the A(,2)B(,1a) subunit. A(,2) had 278 residues and a molecular weight of 31,600 (+OR-) 100. This was considerably less than previously estimated (37,000) by SDS-electrophoresis. B(,1a) contained 180 residues and had a weight of 19,900 + 100. Sequence heterogeneity was detected in both chains and indicated that each was a member of a family of closely related proteins. Sequence homology between chains was not evident. The glycinin composition of cultivar Raiden was determined by purifying and identifying the individual polypeptides. A(,4), A(,5) and B(,3) were present at very low levels, if at all, compared with CX635. This was consistent with the hypothesis that B(,3) was noncovalently associated with A(,4) in addition to being covalently linked to A(,5). Several wild soybean species were analyzed by SDS-electrophoresis. Their glycinin and (beta)-conglycinin compositions varied considerably and were dissimilar to those of G. max. The glycinin fraction of one species was purified and characterized. It contained two groups of acidic polypeptides with NH(,2)-terminal sequences similar to those found in G. max. One group was considerably larger (M(,r) (TURNEQ) 50,000) than previously reported for any glycinin polypeptides. The basic polypeptides of this species had similar, but not identical, molecular weights and NH(,2)-terminal sequences compared with CX635.
Degree
Ph.D.
Subject Area
Biology
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