与辅因子及特定DNA序列结合的转录调节因子TraR的结晶及初步X射线衍射研究。

Crystallization and preliminary X-ray diffraction studies of the transcriptional regulator TraR bound to its cofactor and to a specific DNA sequence.

作者信息

Vannini Alessandro, Volpari Cinzia, Gargioli Cesare, Muraglia Ester, De Francesco Raffaele, Neddermann Petra, Di Marco Stefania

机构信息

Department of Biochemistry, IRBM P. Angeletti, 00040 Pomezia, Rome, Italy.

出版信息

Acta Crystallogr D Biol Crystallogr. 2002 Aug;58(Pt 8):1362-4. doi: 10.1107/s0907444902010028. Epub 2002 Jul 20.

DOI:10.1107/s0907444902010028

PMID:12136156

Abstract

TraR is an Agrobacterium tumefaciens transcriptional regulator which binds the pheromone N-3-oxooctanoyl-L-homoserine lactone (AAI) in response to the bacterial population density. The TraR-AAI complex dimerizes and interacts with a specific 18-base-pair DNA sequence (TraBox), activating promoters containing this site. TraR was overexpressed and purified from Escherichia coli. Crystals of the ternary complex, in which dimeric TraR-AAI is bound to the TraBox sequence, have been obtained by the vapour-diffusion method. The crystals belong to space group P2(1)2(1)2(1), with unit-cell parameters a = 66.99, b = 94.67, c = 209.66 A, with two (TraR-AAI)(2)-TraBox complexes in the asymmetric unit. A three-wavelength MAD data set for the seleno-L-methionine-substituted form has been collected to a resolution of 3 A. 20 of the 24 crystallographically independent selenium sites were located as part of the MAD-phasing process.

摘要

TraR是一种根癌农杆菌转录调节因子，它响应细菌群体密度与信息素N-3-氧代辛酰基-L-高丝氨酸内酯（AAI）结合。TraR-AAI复合物二聚化并与特定的18碱基对DNA序列（TraBox）相互作用，激活含有该位点的启动子。TraR在大肠杆菌中过表达并纯化。通过气相扩散法获得了三元复合物的晶体，其中二聚体TraR-AAI与TraBox序列结合。晶体属于空间群P2(1)2(1)2(1)，晶胞参数a = 66.99，b = 94.67，c = 209.66 Å，不对称单位中有两个(TraR-AAI)(2)-TraBox复合物。已收集到硒代-L-甲硫氨酸取代形式的三波长MAD数据集，分辨率为3 Å。在MAD相位分析过程中，确定了24个晶体学独立硒位点中的20个。