Cloney L P, Bekkaoui D R, Feist G L, Lane W S, Hemmingsen S M
Plant Biotechnology Institute, National Research Council, Saskatoon, Saskatchewan, Canada.
Plant Physiol. 1994 May;105(1):233-41. doi: 10.1104/pp.105.1.233.
Plastid chaperonin-60 protein was purified to apparent homogeneity from Brassica napus using a novel protocol. The purified protein, which migrated as a single species by nondenaturing polyacrylamide gel electrophoresis, contained four polypeptides: three variants of p60cpn60 alpha and p60cpn60 beta. Partial amino acid sequence determination demonstrated that each variant of p60cpn60 alpha is a distinct translation product. During this study, additional chaperonin-60 proteins were purified. These proteins, which were free from contaminating plastid chaperonin-60, were separated into at least two high molecular weight species that were resolved only by nondenaturing polyacrylamide gel electrophoresis. These proteins contained three 60-kD polypeptides. Two of these polypeptides were recognized by existing antisera, whereas the third was not. Partial amino acid sequence data revealed that each of these, including the immunologically distinct polypeptide, is a chaperonin-60 subunit of putative mitochondrial origin. The behavior of chaperonin-60 proteins during blue A Dyematrex chromatography suggests that this matrix may be generally useful for the identification of chaperonin-60 proteins.
利用一种新方法从甘蓝型油菜中纯化出了具有明显均一性的质体伴侣蛋白60。纯化后的蛋白在非变性聚丙烯酰胺凝胶电泳中呈现为单一条带，包含四种多肽：三种p60cpn60α变体和p60cpn60β变体。部分氨基酸序列测定表明，p60cpn60α的每个变体都是一个独特的翻译产物。在这项研究中，还纯化出了其他伴侣蛋白60。这些不含质体伴侣蛋白60污染的蛋白被分离为至少两种仅通过非变性聚丙烯酰胺凝胶电泳才能分辨的高分子量条带。这些蛋白包含三种60-kD多肽。其中两种多肽能被现有抗血清识别，而第三种不能。部分氨基酸序列数据显示，这些蛋白中的每一种，包括免疫上不同的多肽，都是假定线粒体来源的伴侣蛋白60亚基。伴侣蛋白60在蓝色A染料亲和色谱中的行为表明，这种基质可能普遍有助于鉴定伴侣蛋白60。
