Pan C Q, Finkel S E, Cramton S E, Feng J A, Sigman D S, Johnson R C
Molecular Biology Institute, University of California, Los Angeles 90095, USA.
J Mol Biol. 1996 Dec 13;264(4):675-95. doi: 10.1006/jmbi.1996.0669.
The Fis protein from Escherichia coli and Salmonella typhimurium regulates many diverse reactions including recombination, transcription, and replication and is one of the most abundant DNA binding proteins present in the cell under certain physiological conditions. As a specific regulator, Fis binds to discrete sites that are poorly related in primary sequence. Analysis of DNA scission by a collection of Fis conjugates to 1,10-phenanthroline-copper combined with comparative gel electrophoresis has shown that the structures of Fis-DNA complexes are highly variable, displaying overall DNA curvatures that range from < or = 50 degrees to > or = 90 degrees. This variability is primarily determined by differential wrapping of flanking DNA around Fis. By contrast, DNA bending within the core recognition regions appears similar among the binding sites that were analyzed. Flanking DNA contacts by Fis depend on the nucleotide sequence and are mediated by an electrostatic interaction with arginine 71 and a hydrogen bond with asparagine 73, both of which are located outside of the helix-turn-helix DNA binding motif. These contacts strongly influence the kinetics of binding. These data, combined with the crystal structure of Fis, have enabled us to generate new models for Fis-DNA complexes that emphasize the variability in DNA structures within the flanking regions.
来自大肠杆菌和鼠伤寒沙门氏菌的Fis蛋白调控着许多不同的反应，包括重组、转录和复制，并且在某些生理条件下是细胞中含量最丰富的DNA结合蛋白之一。作为一种特异性调节因子，Fis结合到一级序列相关性很差的离散位点上。通过将一系列Fis与1,10 - 菲咯啉 - 铜的缀合物进行DNA切割分析，并结合比较凝胶电泳，结果表明Fis - DNA复合物的结构高度可变，呈现出从≤50度到≥90度不等的整体DNA曲率。这种变异性主要由侧翼DNA围绕Fis的不同缠绕方式决定。相比之下，在分析的结合位点中，核心识别区域内的DNA弯曲似乎相似。Fis与侧翼DNA的接触取决于核苷酸序列，并由与精氨酸71的静电相互作用和与天冬酰胺73的氢键介导，这两者都位于螺旋 - 转角 - 螺旋DNA结合基序之外。这些接触强烈影响结合动力学。这些数据，结合Fis的晶体结构，使我们能够生成Fis - DNA复合物的新模型，该模型强调侧翼区域内DNA结构的变异性。
