Liu T, DeRose E F, Mullen G P
Department of Chemistry, University of Wisconsin at Milwaukee 53211.
Protein Sci. 1994 Aug;3(8):1286-95. doi: 10.1002/pro.5560030815.
The DNA binding domain (DBD) of gamma delta resolvase (residues 141-183) is responsible for the interaction of this site-specific DNA recombinase with consensus site DNA within the gamma delta transposable element in Escherichia coli. Based on chemical-shift comparisons, the proteolytically isolated DBD displays side-chain interactions within a hydrophobic core that are highly similar to those of this domain when part of the intact enzyme (Liu T, Liu DJ, DeRose EF, Mullen GP, 1993, J Biol Chem 268:16309-16315). The structure of the DBD in solution has been determined using restraints obtained from 2-dimensional proton NMR data and is represented by 17 conformers. Experimental restraints included 458 distances based on analysis of nuclear Overhauser effect connectivities, 17 phi and chi 1 torsion angles based on analysis of couplings, and 17 backbone hydrogen bonds determined from NH exchange data. With respect to the computed average structure, these conformers display an RMS deviation of 0.67 A for the heavy backbone atoms and 1.49 A for all heavy atoms within residues 149-180. The DBD consists of 3 alpha-helices comprising residues D149-Q157, S162-T167, and R172-N183. Helix-2 and helix-3 form a backbone fold, which is similar to the canonical helix-turn-helix motif. The conformation of the NH2-terminal residues, G141-R148, appears flexible in solution. A hydrophobic core is formed by side chains donated by essentially all hydrophobic residues within the helices and turns. Helix-1 and helix-3 cross with a right-handed folding topology. The structure is consistent with a mechanism of DNA binding in which contacts are made by the hydrophilic face of helix-3 in the major groove and the amino-terminal arm in the minor groove. This structure represents an important step toward analysis of the mechanism of DNA interaction by gamma delta resolvase and provides initial structure-function comparisons among the divergent DBDs of related resolvases and invertases.
γδ 解离酶的 DNA 结合结构域（第 141 - 183 位氨基酸残基）负责这种位点特异性 DNA 重组酶与大肠杆菌中 γδ 转座元件内共有位点 DNA 的相互作用。基于化学位移比较，经蛋白酶解分离得到的 DNA 结合结构域在疏水核心内显示出侧链相互作用，这与该结构域作为完整酶的一部分时的情况高度相似（刘 T、刘 DJ、德罗塞 EF、马伦 GP，1993 年，《生物化学杂志》268:16309 - 16315）。已利用从二维质子核磁共振数据获得的约束条件确定了溶液中 DNA 结合结构域的结构，其由 17 个构象体表示。实验约束条件包括基于核 Overhauser 效应连接性分析得到的 458 个距离、基于耦合分析得到的 17 个 φ 和 χ1 扭转角，以及根据 NH 交换数据确定的 17 个主链氢键。相对于计算得到的平均结构，这些构象体在第 149 - 180 位氨基酸残基内，重原子主链的均方根偏差为 0.67 Å，所有重原子的均方根偏差为 1.49 Å。DNA 结合结构域由 3 个 α 螺旋组成，分别包含 D149 - Q157、S162 - T167 和 R172 - N183 位氨基酸残基。螺旋 2 和螺旋 3 形成一个主链折叠，类似于典型的螺旋 - 转角 - 螺旋基序。NH2 末端残基 G141 - R148 在溶液中的构象似乎具有灵活性。疏水核心由螺旋和转角内基本上所有疏水残基贡献的侧链形成。螺旋 1 和螺旋 3 以右手折叠拓扑结构交叉。该结构与一种 DNA 结合机制一致，即通过螺旋 3 的亲水面向大沟以及氨基末端臂向小沟进行接触。这一结构代表了朝着分析 γδ 解离酶的 DNA 相互作用机制迈出的重要一步，并在相关解离酶和转化酶不同的 DNA 结合结构域之间提供了初步的结构 - 功能比较。
