Zeidler R, Hobert O, Johannes L, Faulhammer H, Krauss G
Lehrstuhl Biochemie, Universität Bayreuth, Germany.
J Biol Chem. 1993 Sep 25;268(27):20191-7.
We report here the identification and characterization of two novel proteis from Saccharomyces cerevisiae that bind to the T-rich strand of the core consensus autonomously replicating sequence (ARS) in a highly specific manner. The two proteins, 40 and 45 kDa in size, can be distinguished by multiple criteria from each other and from the 65-kDa ssArS-T-binding protein identified recently in our laboratory (Schmidt, A. M. A., Herterich, S. U., and Krauss, G. (1991) EMBO J. 10, 981-985). The specificity of binding is inferred from gel shift and nuclease-footprinting experiments using single-stranded probes containing the core consensus ARS. With a 321-nucleotide single-stranded ARS1 fragment, specific protection of the A and B1 domain against DNase I digestion is observed. Partial amino acid sequencing and enzymatic assays identify the 45-kDa protein as adenylosuccinate synthetase, an enzyme necessary for the de novo synthesis of adenylate.
我们在此报告从酿酒酵母中鉴定和表征出的两种新蛋白质，它们以高度特异性的方式与核心共有自主复制序列（ARS）的富含T的链结合。这两种蛋白质的大小分别为40 kDa和45 kDa，可以通过多种标准相互区分，也可与我们实验室最近鉴定出的65 kDa的ssArS - T结合蛋白区分开来（施密特，A.M.A.，赫特里希，S.U.，和克劳斯，G.（1991年）《欧洲分子生物学组织杂志》10，981 - 985）。结合的特异性是通过使用含有核心共有ARS的单链探针进行凝胶迁移和核酸酶足迹实验推断出来的。对于一个321个核苷酸的单链ARS1片段，观察到A和B1结构域对DNase I消化具有特异性保护作用。部分氨基酸测序和酶活性测定确定45 kDa的蛋白质为腺苷酸琥珀酸合成酶，这是一种从头合成腺苷酸所必需的酶。
