New crystal structures of human glutathione transferase A1-1 shed light on glutathione binding and the conformation of the C-terminal helix
New crystal structures of human glutathione transferase A1-1 shed light on glutathione binding and the conformation of the C-terminal helix
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DOI：
10.1107/s0907444905039296
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发表时间：
2006-02-01
影响因子：
2.2
通讯作者：
Kleywegt, GJ
中科院分区：
文献类型：
--
作者：
Grahn, E;Novotny, M;Kleywegt, GJ
人类谷胱甘肽转移酶A1-1是一种被充分研究的酶，但尽管有丰富的结构和生化数据，但其催化功能的许多方面仍然知之甚少。在这里，描述了这种酶的五个新的晶体结构，提供了一些见解。首先，首次在2.0埃分辨率下测定了野生型人酶与谷胱甘肽复合物的结构。这表明谷胱甘肽在G位点的结合方式与其他结构中底物类似物的谷胱甘肽部分非常相似，而且谷胱甘肽单独结合足以稳定蛋白质的c端螺旋。其次，我们已经研究了与脱羧谷胱甘肽缀合物的复合物，已知该复合物可显着降低酶的活性。人类谷胱甘肽转移酶A1-1的T68E突变体恢复了一些与脱羧谷胱甘肽一起失去的活性，但我们对这种突变体的结构表明，早期对这种现象的解释都不太可能是正确的。第三，偶然地，载脂蛋白结构也揭示了关键的c端区域的构象，这在所有以前的载脂蛋白结构中都是无序的。c端即使在载脂蛋白状态下也能呈有序的螺旋状结构，但有很强的解旋倾向。两种单体载子态的c端结构不同，表明协同作用可能在酶的活性中起作用。
Human glutathione transferase A1-1 is a well studied enzyme, but despite a wealth of structural and biochemical data a number of aspects of its catalytic function are still poorly understood. Here, five new crystal structures of this enzyme are described that provide several insights. Firstly, the structure of a complex of the wild-type human enzyme with glutathione was determined for the first time at 2.0 angstrom resolution. This reveals that glutathione binds in the G site in a very similar fashion as the glutathione portion of substrate analogues in other structures and also that glutathione binding alone is sufficient to stabilize the C-terminal helix of the protein. Secondly, we have studied the complex with a decarboxylated glutathione conjugate that is known to dramatically decrease the activity of the enzyme. The T68E mutant of human glutathione transferase A1-1 recovers some of the activity that is lost with the decarboxylated glutathione, but our structures of this mutant show that none of the earlier explanations of this phenomenon are likely to be correct. Thirdly, and serendipitously, the apo structures also reveal the conformation of the crucial C-terminal region that is disordered in all previous apo structures. The C-terminal region can adopt an ordered helix-like structure even in the apo state, but shows a strong tendency to unwind. Different conformations of the C-terminal regions were observed in the apo states of the two monomers, which suggests that cooperativity could play a role in the activity of the enzyme.