Crystallization and structural analysis of three transcriptional regulators from eubacteria and archaea: Aes. Mlc and TrmB

我们建议结晶并确定与这些调节因子相互作用的原核转录因子和蛋白质的三维结构，这些调节因子控制它们的活性并将这种控制与代谢信号转导链偶联。第一个蛋白质是来自大肠杆菌的Aes。它是一种具有乙酰酯酶活性的酶，与激素敏感脂肪酶家族同源。对这种酶的兴趣是基于它与大肠杆菌麦芽糖系统的控制调节因子MalT的相互作用。第二种转录调节因子是Mlc，它是一种全局转录抑制因子，控制麦芽t基因的表达，此外还有一些基因编码一些特定蛋白质（葡萄糖和甘露糖）和大肠杆菌磷酸转移酶系统（PTS）的所有一般酶。对Mlc的兴趣来自于其通过隔离到PtsG转蛋白的不寻常的失活模式。我们计划确定Mlc的结构，以及Mlc与PtsG可溶性EIIB结构域的配合物的结构。第三种转录调节因子是TrmB，这是一种糖特异性转录抑制因子，控制着嗜热古细菌热球菌（Thermococcus litoralis）中海藻糖和麦芽糖的高亲和力和结合蛋白依赖性ABC转蛋白的编码基因。对这种调节剂的兴趣来自于这样一个事实，即它是第一个确定的古细菌的糖特异性调节剂，作用于真核生物相关的转录机制。这一建议的目标是旨在理解一种新型的转录调控，其中调节剂由直接的蛋白质-蛋白质相互作用控制。

We propose to crystallize and determine the three-dimensional structure of procaryotic transcription factors and proteins that are interacting with these regulators, that control their activity and couple this control to metabolic signal transduction chains. The frist protein is Aes from E. coli. It is an enzyme with acetyl esterase activity that exhibits homology to the family of hormone-sensitive lipases. The interest in this enzyme is based on its demonstrated interaction with MalT, the contral regulator of the E. coli maltose system. The second transcription regulator is Mlc, a global transcriptional repressor controlling the expression of the malT gene and in addition a number of genes encoding some specific proteins (for glucose and mannose) and all general enzymes of the E. coli phosphotransferase system (PTS). The interest in Mlc comes from its unusual mode of inactivation by sequestration to the PtsG transproter. We plan to determine the structure of Mlc als well as of the complex of Mlc with the soluble EIIB domain of PtsG. The third transcription regulator is TrmB, a sugar-specific transcription repressor controlling the genes encoding a high affinity and binding protein-dependent ABC transproter for trehalose and maltose in the hyperthermophilic archaeon Thermococcus litoralis. The interest in this regulator comes from the fact that is ist the first identified sugar specific regulator of archaea acting on a transcription machinery of eucaryotic relateness. The goal of this proposal is aimed at the understanding of a novel type of transcriptional regulation in which the regulator is controlled by direct protein-protein interaction.