No abstract

The molecular weight and subunit composition ofglutelin, the major storage protein of rice, in the major type of protein bodies of developing rice seeds was examined by gradient and twodimensional sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). Glutelin in the protein body was the assembled from heterogeneous subunits, and the molecular weights were estimated to be 64, 140, 240, 320, 380, and 500k by gradient SDS-PAGE.High molecular weight proteins (larger than 2,000 k) were also observed. The two-dimensional SDS-PAGE under reduced conditions showed, that the glutelin in the protein body was composed of two groups of polypeptides, 22~23 and 37~39k, bound by disulfide linkages.

In a study of the relationship between the dynamic changes in cellular organelles and biosynthesis of soybean storage proteins, we found that as the protein componentsaccumulated, the a and a'-subunits of 7S globulin appeared first, followed successively by the acidic and basic subunits of US globulin and finally by the^-subunit of 7S globulin. Labeling experiments using tissue slices showed that the radioactivities of (3H)leucine and (14C)7V-acetyl-D-glycosamine were incorporated into the trichloroacetic acid insoluble fraction and that (14C)7V-acetyl-D-glycosamine was specifically incorporated into 7S. Soybeans have been an important protein source in Japan for centuries. They contain

Rice glutelin occupies about 80%of protein in grains and is accumulated in one type of protein bodies (PB-II) in starchy endosperm of grain.X) Polypeptides of the glutelin are composed of two groups of polypeptides,1>2) and their molecular weights are around 22k and 38k, which are the basic and acidic subunits of glutelin,2) respectively. Although the protein was subdivided into isomers of different molecular weight,3) the subunit composition of glutelin isomers was not clear. In this communication, we fractionated the isomers of rice glutelin by gel filtration and examined their subunit components. Protein bodies were isolated from 100g of milled rice endosperm (Oryza sativa cv. Koshihikari) by the aqueous two-phase system described elsewhere1} except that the concentration ofDEAEdextran was 0. 1 %. After defatting with excess cold acetone (-20°C), glutelin in the protein body fraction was purified by removing other proteins with sequential extraction using 1M NaCl and 55% npropanol.4)The resultant slurry was a crude glutelin fraction that contained about 0.1 g of protein.The glutelin fraction thus obtained was dissolved in 5 ml of 50mMphosphate buffer (pH 6.5) containing 5% SDS, SDS-PAGE was done by the method of Laemmli5) with or without 2-ME in a slab gel. SDS-PAGE profiles in the absence of 2-ME of fractionated rice glutelins are shown in lanes of a-c in Fig. 1. Molecular sizes were assigned to that in PB-II as described elesewhere.3) Their subunit compositions were examined by SDS-PAGE in the presence of 2-ME. Although all proteins were composed of basic(B) and acidic(A) subunits of glutelin, the main polypeptides found in the 64 kD protein fraction (lane d) migrated faster than those in the 130kD and high-molecular-mass proteins (lanes e and f, respectively in Fig. 1). We tentatively numbered them from smaller one to larger one as shown in Fig. 1. The 64-kD protein are basically formed from Bx and Ax subunits judging from the mobilities of these polypeptides. In fractions of the 130-kD protein and the high-molecular-mass proteins, B2, A2, and A3 subunits were predominantly observed. These results showed that 64-kD glutelin was formed from one molecule of Bx and one molecule of Ax. In contrast 130-kD and highmolecular-mass proteins were formed by the combination of two molecule each ofB2 and one molecules ofA2 and A3. Peptic mapping of proteins were done after BrCN degradation^of proteins. After 0.4ml of fractionated proteins (A2so=0.l) was lyophylized, 0.5ml of cyanogen bromide (0.1 g/ml) in 0.1 n HC1 was added to each sample and incubated at room temperature for 24hr in the dark. The reaction was stopped by adding 10 volumes of water, then the mixture was lyophylized. Fifty jul of the sample solution was examined by SDS-PAGE with a 15% polyacrylamide slab gel and silver staining method.7) The electrophoregram showed that peptic maps of the 130-kD protein fraction were the same as that of high-molecularmass proteins and it was different from that of the 64-kD protein (Fig. 2). These results indicated ...