The homologous recombination system of phage lambda. Pairing activities of beta protein.
The homologous recombination system of phage lambda. Pairing activities of beta protein.
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DOI：
10.1016/s0021-9258(17)38416-8
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发表时间：
1986-06
期刊：
影响因子：
--
通讯作者：
K. Muniyappa;C. Radding
中科院分区：
文献类型：
--
作者：
K. Muniyappa;C. Radding
噬菌体λ的红色基因指定两种蛋白质，核酸外切酶和β蛋白，这是其在recA-细胞中的一般遗传重组所必需的。这些蛋白质似乎在体内以等摩尔复合物的形式存在。此外，β蛋白与另一种多肽（可能是噬菌体来源的）形成Mr为70，000的复合物。70-kDa蛋白似乎既不是核酸外切酶或β蛋白的前体也不是聚集形式，因为针对后两种蛋白的抗体在Ouchterlony双扩散分析中未能与70-kDa蛋白反应。β蛋白促进互补链的Mg 2+依赖性复性（Kmiec，E.，和Holloman，W. K.（1981）J.Biol.Chem.256，12636-12639）。为了寻找β蛋白的其他配对活性，我们开发了纯化方法以使其脱离相关的核酸外切酶。不含核酸外切酶的β蛋白似乎不能引起单链与双链DNA的配对;然而，与大肠杆菌单链结合蛋白（SSB）一样，β蛋白刺激recA蛋白从线性双链DNA和同源环状单链形成接合分子。与recA蛋白类似，但与SSB不同，β蛋白促进噬菌体λ DNA互补单链末端的连接。β蛋白特异性地保护单链DNA免受胰腺DNA酶的消化。β蛋白催化的复性半衰期与DNA浓度无关，这与SSB促进的复性和自发复性不同，后者是二级反应。因此，β蛋白在将单链DNA分子聚集在一起的能力方面类似于recA蛋白，并且在减少单链DNA二级结构的能力方面类似于SSB。
The red genes of phage lambda specify two proteins, exonuclease and beta protein, which are essential for its general genetic recombination in recA- cells. These proteins seem to occur in vivo as an equimolar complex. In addition, beta protein forms a complex with another polypeptide, probably of phage origin, of Mr 70,000. The 70-kDa protein appears to be neither a precursor nor an aggregated form of either exonuclease or beta protein, since antibodies directed against the latter two proteins failed to react with 70-kDa protein on Ouchterlony double diffusion analysis. beta protein promotes Mg2+-dependent renaturation of complementary strands (Kmiec, E., and Holloman, W. K. (1981) J. Biol. Chem. 256, 12636-12639). To look for other pairing activities of beta protein, we developed methods of purification to free it of associated exonuclease. Exonuclease-free beta protein appeared unable to cause the pairing of a single strand with duplex DNA; however, like Escherichia coli single strand binding protein (SSB), beta protein stimulated formation of joint molecules by recA protein from linear duplex DNA and homologous circular single strands. Like recA protein, but unlike SSB, beta protein promoted the joining of the complementary single-stranded ends of phage lambda DNA. beta protein specifically protected single-stranded DNA from digestion by pancreatic DNase. The half-time for renaturation catalyzed by beta protein was independent of DNA concentration, unlike renaturation promoted by SSB and spontaneous renaturation, which are second order reactions. Thus, beta protein resembles recA protein in its ability to bring single-stranded DNA molecules together and resembles SSB in its ability to reduce secondary structure in single-stranded DNA.