Stupperich E, Aulkemeyer P, Eckerskorn C
Abteilung Angewandte Mikrobiologie, Universität Ulm, Federal Republic of Germany.
Arch Microbiol. 1992;158(5):370-3. doi: 10.1007/BF00245367.
The major cobamide-containing protein from methanol-utilizing Sporomusa ovata was 8-fold enriched to apparent homogeneity. The protein exhibited a molecular mass of 40 kDa and of 38 kDa determined by gel filtration and by SDS-polyacrylamide gel electrophoresis, respectively. This finding indicates a monomeric protein structure. Monospecific polyclonal antisera raised against the protein did not cross react with another cobamide-containing protein from Sporomusa cells. Only the 40 kDa cobamide-containing protein was induced by methanol, since proteins from cells grown on 3,4-dimethoxybenzoate, betaine H2/CO2, or fructose showed faint or no cross reaction. Hence, the 40 kDa cobamide-containing protein is presumably involved in the methyl-transfer reaction of the methanol metabolism. The purified enzyme revealed 1.1 mol of p-cresolyl cobamide per mol of protein, but it lacked of iron-sulfur centers. Remarkably, the cofactor was firmly bound to its protein.
利用甲醇的卵形芽孢杆菌中主要的含钴胺素蛋白被富集了8倍，达到了明显的均一性。通过凝胶过滤和SDS-聚丙烯酰胺凝胶电泳分别测定，该蛋白的分子量为40 kDa和38 kDa。这一发现表明其为单体蛋白结构。针对该蛋白制备的单特异性多克隆抗血清与卵形芽孢杆菌细胞中的另一种含钴胺素蛋白不发生交叉反应。只有40 kDa的含钴胺素蛋白是由甲醇诱导产生的，因为在3,4-二甲氧基苯甲酸、甜菜碱H2/CO2或果糖上生长的细胞中的蛋白显示出微弱的交叉反应或无交叉反应。因此，40 kDa的含钴胺素蛋白可能参与甲醇代谢的甲基转移反应。纯化后的酶每摩尔蛋白含有1.1摩尔对甲酚基钴胺素，但缺乏铁硫中心。值得注意的是，辅因子与蛋白紧密结合。
