Kang H, Lee S G, Cho Y D
Department of Medical Technology, College of Allied Health Science, Korea University, Seoul 136-703, Korea.
Biochem J. 1998 Jun 1;332 ( Pt 2)(Pt 2):467-73. doi: 10.1042/bj3320467.
To identify a polyamine-conjugated protein by the action of transglutaminase in the absence of radiolabelled polyamine, extracts prepared from the leaves and developing soybean seeds were investigated for the specific activity of transglutaminase and the content of free polyamines. We identified the major storage protein, glycinin, as a polyamine-conjugated protein. This was established by the following procedures: (1) immunolocalization with antibody against putrescine prepared in rabbit against putrescine-BSA conjugate; (2) immunocross-reactivity on nitrocellulose transblot of the purified glycinin subunits by using antibody against putrescine; (3) identification of polyamines in acid hydrolysates of purified glycinin; (4) release of polyamines in proteolytic digests through the catalytic action of gamma-glutamylamine cyclotransferase, an enzyme specific for the disassembly of gamma-glutamylamines. The activity of gamma-glutamylamine cyclotransferase was also identified in soybean seeds.
为了在不存在放射性标记多胺的情况下通过转谷氨酰胺酶的作用鉴定多胺缀合蛋白，对从叶片和发育中的大豆种子制备的提取物进行了转谷氨酰胺酶比活性和游离多胺含量的研究。我们鉴定出主要的贮藏蛋白大豆球蛋白是一种多胺缀合蛋白。这是通过以下步骤确定的：（1）用兔抗腐胺 - BSA缀合物制备的抗腐胺抗体进行免疫定位；（2）使用抗腐胺抗体对纯化的大豆球蛋白亚基在硝酸纤维素转印膜上进行免疫交叉反应；（3）鉴定纯化大豆球蛋白酸水解产物中的多胺；（4）通过γ-谷氨酰胺环转移酶（一种特异性拆解γ-谷氨酰胺的酶）的催化作用，在蛋白水解消化物中释放多胺。在大豆种子中也鉴定出了γ-谷氨酰胺环转移酶的活性。
