Three-dimensional structure of N5-carboxyaminoimidazole ribonucleotide synthetase:: A member of the ATP grasp protein superfamily
Three-dimensional structure of N5-carboxyaminoimidazole ribonucleotide synthetase:: A member of the ATP grasp protein superfamily
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DOI：
10.1021/bi991618s
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发表时间：
1999-11-23
期刊：
影响因子：
2.9
通讯作者：
Holden, HM
中科院分区：
文献类型：
--
作者：
Thoden, JB;Kappock, TJ;Holden, HM
大肠杆菌PurK是一种二聚体的N - 5 - 羧基氨基咪唑核糖核苷酸（N - 5 - CAIR）合成酶，它催化5 - 氨基咪唑核糖核苷酸（AIR）、ATP和碳酸氢盐转化为N - 5 - CAIR、ADP和磷酸（Pi）。硫酸根结合的以及MgADP结合的大肠杆菌PurK的结晶分别得到了分辨率为2.1埃和2.5埃的结构。PurK属于C - N连接酶的ATP抓握超家族。PurK的每个亚基由三个结构域（A、B和C）组成。B结构域包含一个灵活的、富含甘氨酸的环（B环，T - 123 - G(130)），它在硫酸根 - PurK结构中是无序的，而在MgADP - PurK结构中变得有序。MgADP像ATP抓握超家族的所有成员一样，楔入B和C结构域之间。这个超家族中的其他酶包含一个保守的Ω环，据推测它与B环相互作用，决定其非核苷酸底物的特异性，并保护由该底物形成的酰基磷酸中间体。PurK包含一个最小的Ω环，没有保守残基。在PurK催化的反应中，羧基磷酸是假定的酰基磷酸中间体。硫酸根离子结合的PurK中的硫酸根与Arg 242和Asn 245的主链酰胺基静电相互作用，Asn 245是C结构域J环的组成部分。这种硫酸根可能揭示了羧基磷酸结合位点的位置。C结构域C末端内的保守残基定义了一个口袋，据推测它与A结构域中的一个N末端链 - 环 - 螺旋基序（P环，G(8) - L - 12）协作结合AIR。根据在PurN和PurE中观察到的类似结合位点以及在嘌呤途径中的其他四种酶PurD和PurT中所推测的情况，P环被推测结合AIR的磷酸基团。
Escherichia coli PurK, a dimeric N-5-carboxyaminoimidazole ribonucleotide (N-5-CAIR) synthetase, catalyzes the conversion of 5-aminoimidazole ribonucleotide (AIR), ATP, and bicarbonate to N-5-CAIR, ADP, and Pi. Crystallization of both a sulfate-liganded and the MgADP-liganded E. coli PurK has resulted in structures at 2.1 and 2.5 Angstrom resolution, respectively. PurK belongs to the ATP grasp superfamily of C-N ligase enzymes, Each subunit of PurK is composed of three domains (A, B, and C). The B domain contains a flexible, glycine-rich loop (B loop, T-123-G(130)) that is disordered in the sulfate-PurK structure and becomes ordered in the MgADP-PurK structure. MgADP is wedged between the B and C domains, as with all members of the ATP grasp superfamily. Other enzymes in this superfamily contain a conserved Omega loop proposed to interact with the B loop, define the specificity of their nonnucleotide substrate, and protect the acyl phosphate intermediate formed from this substrate. PurK contains a minimal Omega loop without conserved residues. In the reaction catalyzed by PurK, carboxyphosphate is the putative acyl phosphate intermediate. The sulfate of the sulfate ion-liganded PurK interacts electrostatically with Arg 242 and the backbone amide group of Asn 245, components of the J loop of the C domain. This sulfate may reveal the location of the carboxyphosphate binding site, Conserved residues within the C-terminus of the C domain define a pocket that is proposed to bind AIR in collaboration with an N-terminal strand loop helix motif in the A domain (P loop, G(8)-L-12) The P loop is proposed to bind the phosphate of AIR on the basis of similar binding sites observed in PurN and PurE and proposed in PurD and PurT, four other enzymes in the purine pathway.