Tan F L, Shi Q L, Wang L X, Qi Z W
Sci Sin B. 1984 Sep;27(9):918-25.
Two peptide chains A1 and A2 of the Lys active fragment, linked via a couple of inter-disulfide bonds, could be separated from each other after reduction with dithiothreitol and gel filtration on Sephadex G-25. Reoxidation of the reduced peptide chain A1 resulted in recovering the inhibitory activity with 25% yield, based on the original activity of the Lys fragment. The A1 active fragment was further purified by affinity chromatography with immobilized trypsin. Sephadex G-25 gel filtration produced two forms of the A1 active fragment, the major fraction being a monomer and the minor one being a dimer with lower activity. The results obtained offered evidence of the evolution of mung bean inhibitor from an ancestral single-headed inhibitor by fused gene duplication with A2 as a connecting peptide. The CD spectra of the Lys fragment and the reoxidized peptide chain A1 were also compared.
赖氨酸活性片段的两条肽链A1和A2通过一对链间二硫键相连，用二硫苏糖醇还原并在葡聚糖凝胶G-25上进行凝胶过滤后，它们可以彼此分离。还原后的肽链A1再氧化后，基于赖氨酸片段的原始活性，以25%的产率恢复了抑制活性。A1活性片段通过固定化胰蛋白酶亲和层析进一步纯化。葡聚糖凝胶G-25凝胶过滤产生了两种形式的A1活性片段，主要部分是单体，次要部分是活性较低的二聚体。所得结果为绿豆抑制剂从祖先的单头抑制剂通过融合基因复制进化而来提供了证据，其中A2作为连接肽。还比较了赖氨酸片段和再氧化肽链A1的圆二色谱。
