Crystal structures of unbound and aminooxyacetate-bound Escherichia coli γ-aminobutyrate aminotransferase
Crystal structures of unbound and aminooxyacetate-bound Escherichia coli γ-aminobutyrate aminotransferase
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DOI：
10.1021/bi049218e
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发表时间：
2004-08-31
期刊：
影响因子：
2.9
通讯作者：
Toney, MD
中科院分区：
文献类型：
--
作者：
Liu, WS;Peterson, PE;Toney, MD
本文报道了大肠杆菌γ-氨基丁酸氨基转移酶的晶体结构。该酶从硫酸铵溶液中以P3（2）21空间群结晶，在不对称单元中具有四聚体。收集衍射数据，对于未配体的酶，分辨率为2.4埃，对于氨氧基乙酸盐复合物，分辨率为1.9埃。该酶的总体结构与其他氨基转移酶亚组11酶的结构相似。γ-氨基丁酸转氨酶作用于伯胺底物的能力在第一半反应中的（γ-氨基丁酸）和第二半反应中的α-氨基酸之间的相互作用被认为是通过Glu 211的存在而实现的，Glu 211的侧链羧酸酯在与伯胺半反应中的Arg 398的相互作用和与α-氨基酸半反应中的替代结合位点之间交替，其中Arg 398结合底物α-羧酸。对底物侧链上的羧酸基团的特异性主要是由于Arg 141的存在，但也需要相对于结构同源的酶二烷基甘氨酸脱羧酶的大量局部主链重排，所述酶对小烷基侧链具有特异性。在细菌酶中没有发现如在猪酶中发现的铁-硫簇[Storici，P.，De Biase，D.，Bossa，F.，Bruno，S.，Mozzarelli，A.，佩内夫角，西尔弗曼河B.，和Schirmer，T.（2004）J.Biol.Chem.279，363-73.]。氨氧乙酸的结合导致活性位点结构的显著小的变化，并且没有观察到大的结构域运动。活性部位的结构与猪γ-氨基丁酸转氨酶和二烷基甘氨酸脱羧酶进行了比较讨论。
The X-ray crystal structures of Escherichia coli gamma-aminobutyrate aminotransferase unbound and bound to the inhibitor aminooxyacetate are reported. The enzyme crystallizes from ammonium sulfate solutions in the P3(2)21 space group with a tetramer in the asymmetric unit. Diffraction data were collected to 2.4 Angstrom resolution for the unliganded enzyme and 1.9 Angstrom resolution for the aminooxyacetate complex. The overall structure of the enzyme is similar to those of other aminotransferase subgroup 11 enzymes. The ability of gamma-aminobutyrate aminotransferase to act on primary amine substrates (gamma-aminobutyrate) in the first half-reaction and a-amino acids in the second is proposed to be enabled by the presence of Glu211, whose side chain carboxylate alternates between interactions with Arg398 in the primary amine half-reaction and an alternative binding site in the alpha-amino acid half-reaction, in which Arg398 binds the substrate alpha-carboxylate. The specificity for a carboxylate group on the substrate side chain is due primarily to the presence of Arg141, but also requires substantial local main chain rearrangements relative to the structurally homologous enzyme dialkylglycine decarboxylase, which is specific for small alkyl side chains. No iron-sulfur cluster is found in the bacterial enzyme as was found in the pig enzyme [Storici, P., De Biase, D., Bossa, F., Bruno, S., Mozzarelli, A., Peneff, C., Silverman, R. B., and Schirmer, T. (2004) J. Biol. Chem. 279, 363-73.]. The binding of aminooxyacetate causes remarkably small changes in the active site structure, and no large domain movements are observed. Active site structure comparisons with pig gamma-aminobutyrate aminotransferase and dialkylglycine decarboxylase are discussed.