Smirnova Irina A, Dian Cyril, Leonard Gordon A, McSweeney Seán, Birse Darcy, Brzezinski Peter
Department of Biochemistry and Biophysics, Arrhenius Laboratories for Natural Sciences, Stockholm University, SE-106 91 Stockholm, Sweden.
J Mol Biol. 2004 Jul 9;340(3):405-18. doi: 10.1016/j.jmb.2004.04.071.
The transcriptional regulator DntR, a member of the LysR family, is a central element in a prototype bacterial cell-based biosensor for the detection of hazardous contamination of soil and groundwater by dinitrotoluenes. To optimise the sensitivity of the biosensor for such compounds we have chosen a rational design of the inducer-binding cavity based on knowledge of the three-dimensional structure of DntR. We report two crystal structures of DntR with acetate (resolution 2.6 angstroms) and thiocyanate (resolution 2.3 angstroms), respectively, occupying the inducer-binding cavity. These structures allow for the construction of models of DntR in complex with salicylate (Kd approximately or = 4 microM) and 2,4-dinitrotoluene that provide a basis for the design of mutant DntR with enhanced specificity for dinitrotoluenes. In both crystal structures DntR crystallises as a homodimer with a "head-to-tail" arrangement of monomers in the asymmetric unit. Analysis of the crystal structure has allowed the building of a full-length model of DntR in its biologically active homotetrameric form consisting of two "head-to-head" dimers. The implications of this model for the mechanism of transcription regulation by LysR proteins are discussed.
转录调节因子DntR是LysR家族的成员，是一种基于原型细菌细胞的生物传感器的核心元件，用于检测土壤和地下水中二硝基甲苯的有害污染。为了优化生物传感器对此类化合物的灵敏度，我们基于DntR的三维结构知识，对诱导物结合腔进行了合理设计。我们分别报道了DntR与乙酸盐（分辨率2.6埃）和硫氰酸盐（分辨率2.3埃）的两种晶体结构，它们分别占据诱导物结合腔。这些结构使得构建DntR与水杨酸盐（解离常数约为或 = 4 microM）和2,4 - 二硝基甲苯复合物的模型成为可能，为设计对二硝基甲苯具有更高特异性的突变型DntR提供了基础。在这两种晶体结构中，DntR均以同型二聚体形式结晶，不对称单元中的单体呈“头对尾”排列。对晶体结构的分析使得构建DntR以其具有生物活性的同型四聚体形式（由两个“头对头”二聚体组成）的全长模型成为可能。本文讨论了该模型对LysR蛋白转录调控机制的影响。
