我国大豆种子中球蛋白2S组分的分离纯化及部分性质的研究

The Seperation, Purification and Some Properties of 2S Fraction of Soybean Seed Globulins

根据大豆种子球蛋白和清蛋白溶解性不同的原理,分离出我国红丰3号大豆种子球蛋白2S粗制剂,再用SephadexG-100柱层析对其进行纯化。纯化后的球蛋白表现SDS-聚丙烯酰胺凝胶电泳非均一性。对这样纯化过的2s球蛋白进行DEAE-纤维素离子交换柱层析分离,用0.1—0.5mol/L pH7.6磷酸盐缓冲液进行线性梯度洗脱,得到四个洗脱峰,每个峰都获得了PAGE单一条带。四个组分分别命名为SⅡP_1,SⅡP_2,SⅡP_3和SⅡP_4。然后对这四种蛋白质的某些性质进行了研究,结果表明四者的分子量按以上顺序分别为22800,21500,19200和17800。所含残基数分别为191,179,163和147个。SⅡP_1,SⅡP_2和SⅡP_3三者的沉降系数(S_(20),w)分别为2.1S,1.9S和1.8S。N-末端分析表明这四种蛋白质的N-末端均为天门冬氨酸.还发现SⅡP_2具有能抑制α-胰凝乳蛋白酶的活性。本实验所提纯的这个抑制剂的一个ATEE(N-乙酰-L-酪氨酸乙酯)单位为0.4μg抑制剂蛋白(仅指对α-chymotrypsin发生作用)。α-chymotrypsin与此抑制剂相互作用时的摩尔数比初步判断为E/I=2/1。

A crude 2 S fraction was separated from soybean seed globulins and purified by sephadex G-100 column chromatography. The purified 2S globulin gives several bands on SDS-PAGE. Four peaks were eluted when the purified 2 S globulin was further separated on DEAE-Cellulose ion-exchange column by ionic strength gradient elution of from 0.1moL/L to 0.5mol/L. Each of the four components is homogeneous in the patterns of polyacrylamidegel electrophoresis. The four components were nominated SⅡP1, SⅡP2,SⅡP3 and SⅡP4 respectively according to their eluted order. Then some properties were studied. The results showed that the M.W. s of the four proteins are 22 800, 21 500, 19 200 and 17800 respectively. The residue numbers of the four are, SⅡP1 = 191,SⅡP2 = 179,SⅡP3 = 163 and SⅡP4 = 147. The sedimentation coefficent (S20,w) of SⅡP1, SⅡP2 and SⅡP3 were 2.1S, 1.9S and 1.8S. The terminal analysis showed that the same N-terminal amino acid Asp. was found in all these four proteins. It was also found that only SⅡP2 has the activity of inhibiting a-chymotrypsin with E/I ratio (refer to α-chymotrypsin/ SⅡP2) of 2/1.