Perrino F W, Meyer R R, Bobst A M, Rein D C
Department of Biological Sciences, University of Cincinnati, Ohio 45221.
J Biol Chem. 1988 Aug 25;263(24):11833-9.
A single-stranded DNA-binding protein (SSB) affinity column was prepared by optimizing the coupling of Escherichia coli single-stranded DNA-binding protein to Affi-Gel 10. The bound SSB retained its ability to specifically bind single-stranded DNA. When nuclease-treated cell extracts were incubated with the SSB beads overnight at 4 degrees C, a major protein of Mr = 25,000 was bound. At shorter incubation times, two additional proteins of Mr = 32,000 and 36,000 were also detected. In the absence of nuclease treatment, eight additional proteins ranging from Mr = 14,000 to 160,000 also bound to the affinity column. The major Mr = 25,000 protein has been shown to be a folded chromosome-associated protein. Its binding to SSB is strongly enhanced by the addition of DNA polymerase III or DNA polymerase III holoenzyme.
通过优化大肠杆菌单链DNA结合蛋白与Affi-Gel 10的偶联制备了单链DNA结合蛋白（SSB）亲和柱。结合的SSB保留了其特异性结合单链DNA的能力。当用核酸酶处理的细胞提取物在4℃下与SSB珠一起孵育过夜时，一种分子量为25,000的主要蛋白质被结合。在较短的孵育时间下，还检测到另外两种分子量分别为32,000和36,000的蛋白质。在没有核酸酶处理的情况下，另外八种分子量从14,000到160,000的蛋白质也与亲和柱结合。已证明分子量为25,000的主要蛋白质是一种折叠的染色体相关蛋白。添加DNA聚合酶III或DNA聚合酶III全酶可强烈增强其与SSB的结合。
