A pseudo-Michaelis quaternary complex in the reverse reaction of a ligase, structure of Escherichia coli B glutathione synthetase complexed with ADP, glutathione; and sulfate at 2.0 angstrom resolution
A pseudo-Michaelis quaternary complex in the reverse reaction of a ligase, structure of Escherichia coli B glutathione synthetase complexed with ADP, glutathione; and sulfate at 2.0 angstrom resolution
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DOI：
10.1021/bi9605245
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发表时间：
1996-09-17
期刊：
影响因子：
2.9
通讯作者：
Oda, JI
中科院分区：
文献类型：
--
作者：
Hara, T;Kato, H;Oda, JI
在2.0埃分辨率下测定了大肠杆菌B与ADP、谷胱甘肽和硫酸盐形成的谷胱甘肽合成酶的晶体结构。考虑到硫酸盐和磷酸盐的化学相似性，这种四元络合物结构代表了反向反应中的假酶-底物络合物，从而使我们能够了解大肠杆菌谷胱甘肽合成酶的活性部位结构。两个镁离子与ADP的α-和β-磷酸基团以及硫酸盐离子中的氧原子配位。在无配体或二元和三元络合物结构中看不到的柔性环被固定在四元络合物中。较大的柔性环(Ile226-Arg241)包括3(10)-螺旋的一个转角，该螺旋组成GSH的甘氨酸部分的结合部位。小环(Gly164-Gly167)参与核苷酸结合，并充当磷酸夹持器。Arg210和Arg225的侧链与ADP的硫酸盐离子和β-磷酸部分相互作用。Arg 210可能以正向反应的底物结合形式与C末端的伽马-谷氨酰半胱氨酸的羧酸盐相互作用。活性部位(Lys125和Lys160)上的其他正电荷残基参与核苷酸结合，将磷酸基团引导到正确的催化位置，讨论了底物结合形式的活性部位结构的功能方面。
The crystal structure of glutathione synthetase from Escherichia coli B complexed with ADP, glutathione, and sulfate has been determined at 2.0 Angstrom resolution. Concerning the chemical similarity of sulfate and phosphate, this quaternary complex structure represents a pseudo enzyme-substrate complex in the reverse reaction and consequently allows us to understand the active site architecture of the E. coli glutathione synthetase. Two Mg2+ ions are coordinated with oxygen atoms from the alpha- and beta-phosphate groups of ADP and from the sulfate ion. The flexible loops, invisible in the unliganded or the binary and ternary complex structures, are fixed in the quaternary complex. The larger flexible loop (Ile226-Arg241) includes one turn of a 3(10)-helix that comprises the binding site of the glycine moiety of GSH. The small loop (Gly164-Gly167) is involved in nucleotide binding and acts as a phosphate gripper. The side chains of Arg210 and Arg225 interact with the sulfate ion and the beta-phosphate moiety of ADP. Arg 210 is likely to interact with the carboxylate of the C-terminal gamma-glutamylcysteine in the substrate-binding form of the forward reaction. Other positively charged residues in the active site (Lys125 and Lys160) are involved in nucleotide binding, directing the phosphate groups to the right position for catalysis, Functional aspects of the active site architecture in the substrate-binding form are discussed.