Structure and mechanism of E. coli RNA 2',3'-cyclic phosphodiesterase.
Structure and mechanism of E. coli RNA 2',3'-cyclic phosphodiesterase.
复制标题
DOI：
10.1261/rna.046797.114
复制
发表时间：
2014-11
期刊：
影响因子：
--
通讯作者：
Shuman S
中科院分区：
文献类型：
--
作者：
Remus BS;Jacewicz A;Shuman S
2H(2-组氨酸)磷酸酯酶广泛分布于生命的各个领域，参与多种RNA和核苷酸交易，包括环磷酸盐的酯交换和水解酶。本文报道了大肠杆菌2H蛋白YapD的生化和结构特征，该蛋白最初被鉴定为RNA2‘，5’-磷酸二酯的可逆酯交换“核酸酶/连接酶”。我们发现YapD是一种“末端修复”的环磷酸二酯酶(CPDase)，它将含有2‘，3’-环磷酸二酯的Horna&Gt；p底物水解成末端带有2‘-磷酸单酯的HORNAp产物，而不伴随末端连接。因此，我们将该酶重新命名为ThpR(作用于RNA的2-组氨酸2‘，3’-环磷酸二酯酶)。在与2‘-AMP形成的产物络合物中，ThpR的2.0ä晶体结构突出了扩展的含有组氨酸的基序43HxTxxF48和125HxTxxR130在CPDase反应中的作用。His43-Nε与核糖O‘离开基团形成氢键，从而暗示His43是一种普通的酸催化剂。His125-Nε配位AMP2‘-磷酸的O1P氧(从几何角度推断来自攻击水的亲核试剂)，指出His125是一般的碱催化剂。Arg130与AMP2‘-磷酸发生双齿接触，提示其在过渡态稳定中发挥作用。与这些推论一致的是，将His43、His125或Arg130改为丙氨酸会降低ThpR的CPDase活性。Phe48在腺嘌呤碱基上形成π-π堆栈。将Phe28突变为丙氨酸会使CPDase减慢一个数量级。ThpR的三级结构和扩展的活性位点基序在细菌和古生菌的2H酶亚家族中保守。
2H (two-histidine) phosphoesterase enzymes are distributed widely in all domains of life and are implicated in diverse RNA and nucleotide transactions, including the transesterification and hydrolysis of cyclic phosphates. Here we report a biochemical and structural characterization of the Escherichia coli 2H protein YapD, which was identified originally as a reversible transesterifying “nuclease/ligase” at RNA 2′,5′-phosphodiesters. We find that YapD is an “end healing” cyclic phosphodiesterase (CPDase) enzyme that hydrolyzes an HORNA>p substrate with a 2′,3′-cyclic phosphodiester to a HORNAp product with a 2′-phosphomonoester terminus, without concomitant end joining. Thus we rename this enzyme ThpR (two-histidine 2′,3′-cyclic phosphodiesterase acting on RNA). The 2.0 Å crystal structure of ThpR in a product complex with 2′-AMP highlights the roles of extended histidine-containing motifs 43HxTxxF48 and 125HxTxxR130 in the CPDase reaction. His43-Nε makes a hydrogen bond with the ribose O3′ leaving group, thereby implicating His43 as a general acid catalyst. His125-Nε coordinates the O1P oxygen of the AMP 2′-phosphate (inferred from geometry to derive from the attacking water nucleophile), pointing to His125 as a general base catalyst. Arg130 makes bidentate contact with the AMP 2′-phosphate, suggesting a role in transition-state stabilization. Consistent with these inferences, changing His43, His125, or Arg130 to alanine effaced the CPDase activity of ThpR. Phe48 makes a π–π stack on the adenine nucleobase. Mutating Phe28 to alanine slowed the CPDase by an order of magnitude. The tertiary structure and extended active site motifs of ThpR are conserved in a subfamily of bacterial and archaeal 2H enzymes.
登录
查看更多内容
影响因子：
2.9
作者：
Li, Dan;Liu, Cong;Su, Xiao-Dong
通讯作者：
Su, Xiao-Dong
DOI：
10.1107/s0907444904019158
发表时间：
2004-12-01
影响因子：
2.2
作者：
Emsley, P;Cowtan, K
通讯作者：
Cowtan, K
影响因子：
4.5
作者：
Remus, Barbara S.;Shuman, Stewart
通讯作者：
Shuman, Stewart
影响因子：
20.3
作者：
Hilcenko, Christine;Simpson, Paul J.;Warren, Alan J.
通讯作者：
Warren, Alan J.
DOI：
10.1107/s0907444905017841
发表时间：
2005-09-01
影响因子：
2.2
作者：
Rehse, PH;Tahirov, HT
通讯作者：
Tahirov, HT