Sanders G M, Kassavetis G A, Geiduschek E P
Department of Biology, University of California, San Diego, La Jolla 92093-0634, USA.
EMBO J. 1995 Aug 15;14(16):3966-76. doi: 10.1002/j.1460-2075.1995.tb00068.x.
The bacteriophage T4 DNA polymerase accessory proteins confer processivity and high speed on replicative DNA chain elongation: the gene 45 protein, gp45, tracks along DNA and serves as the sliding clamp of the viral DNA polymerase; the gene 44/62 protein complex, gp44/62, is an ATP-dependent loading enzyme that mounts gp45 on DNA. Gp45 also activates T4 late transcription. Transcriptional enhancement by gp45 requires a particular orientation that is imposed by gp44/62 at the DNA loading site. Loading and orienting gp45 on DNA, tracking along DNA and interaction with RNA polymerase have been analyzed by measuring transcriptional activation. The efficiency of loading gp45 at different DNA structures and the resulting transcriptional activation have been compared, and sources of interference with transcriptional activation have been examined. All observations are compatible with a mechanism in which the loading enzyme recognizes the polarity of single-stranded DNA and imposes a corresponding polarity of DNA entry on gp45. Primer-template junctions are the most efficient DNA loading sites for gp45 and can generate very rapid opening at promoters that are located at a distance of > 1 kbp. In contrast, gp45 does not track efficiently across single-stranded DNA.
噬菌体T4 DNA聚合酶辅助蛋白赋予复制性DNA链延伸以持续合成能力和高速度：基因45蛋白（gp45）沿DNA移动，作为病毒DNA聚合酶的滑动夹；基因44/62蛋白复合物（gp44/62）是一种依赖ATP的装载酶，可将gp45装载到DNA上。Gp45还激活T4晚期转录。Gp45介导的转录增强需要特定的方向，该方向由gp44/62在DNA装载位点施加。通过测量转录激活来分析gp45在DNA上的装载、定向、沿DNA移动以及与RNA聚合酶的相互作用。比较了在不同DNA结构上装载gp45的效率以及由此产生的转录激活，并研究了对转录激活的干扰来源。所有观察结果都与一种机制相符，即装载酶识别单链DNA的极性，并将相应的DNA进入极性施加于gp45。引物-模板接头是gp45最有效的DNA装载位点，并且可以在距离>1 kbp的启动子处产生非常快速的开放。相比之下，gp45不能有效地沿单链DNA移动。
