Structural determinants of substrate recognition in the HAD superfamily member D-glycero-D-manno-heptose-1,7-bisphosphate phosphatase (GmhB) .
Structural determinants of substrate recognition in the HAD superfamily member D-glycero-D-manno-heptose-1,7-bisphosphate phosphatase (GmhB) .
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DOI：
10.1021/bi902019q
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发表时间：
2010-02-16
期刊：
影响因子：
2.9
通讯作者：
Allen, Karen N.
中科院分区：
文献类型：
--
作者：
Nguyen, Henry H.;Wang, Liangbing;Huang, Hua;Peisach, Ezra;Dunaway-Mariano, Debra;Allen, Karen N.
卤代链烷酸脱卤酶（HAD）1酶超家族是磷酸水解酶中最大的家族。在HAD成员中，提供支持底物特异性的结合相互作用的结构元件与协调催化的结构元件分开。对于大多数HAD磷酸酶来说，帽结构域在底物识别中发挥作用。然而，对于缺乏帽结构域的HAD磷酸酶，必须有一种替代的底物选择策略。选择HisB亚家族的一种这样的HAD磷酸酶GmhB用于结构-功能分析。本文对E.除测定B的结构外，还测定了大肠杆菌Gmh B的载脂蛋白形式（1.6 μ m分辨率）、与Mg ~（2+）和正磷酸盐的络合物（1.8 μ m分辨率）以及与Mg ~（2+）和D-甘露庚糖-1 β，7-二磷酸盐的络合物（2.2 μ m分辨率）。支气管败血症GmhB与Mg 2+和正磷酸盐结合（1.7 μ M分辨率）。结构表明，在帽结构域的地方，GmhB催化位点是由三个肽插入物或环，包装形成一个凹的，半圆形的表面周围的底物离去基团精心制作。定点突变体的结构指导的动力学分析进行了平行的HisB亚家族内的序列多样性的生物信息学研究，以确定环残基，作为底物识别元件，并区分GmhB从其亚家族对应，组氨醇磷酸磷酸酶结构域的HisB。我们发现，GmhB和磷酸组氨醇磷酸酶结构域使用相同的设计，三个底物识别环插入到帽结构域，但通过选择性残基使用的循环，实现了独特的底物特异性，从而新的生化功能。
The Haloalkanoic Acid Dehalogenase (HAD)1 enzyme superfamily is the largest family of phosphohydrolases. In HAD members, the structural elements that provide the binding interactions that support substrate specificity are separated from those that orchestrate catalysis. For most HAD phosphatases a cap domain functions in substrate recognition. However, for the HAD phosphatases which lack a cap domain, an alternate strategy for substrate selection must be operative. One such HAD phosphatase, GmhB of the HisB subfamily was selected for structure-function analysis. Herein, the X-ray crystallographic structures of E. coli GmhB in the apo form (1.6 Å resolution), complexed with Mg2+ and orthophosphate (1.8 Å resolution), and with Mg2+ and Dglycero-D-manno-heptose-1β,7-bisphosphate (2.2 Å resolution) were determined, in addition to the structure of B. bronchiseptica GmhB bound to Mg2+ and orthophosphate (1.7 Å resolution). The structures show that in place of a cap domain, the GmhB catalytic site is elaborated by three peptide inserts or loops that pack to form a concave, semicircular surface around the substrate leaving group. Structure-guided kinetic analysis of site-directed mutants was carried out in parallel with a bioinformatics study of sequence diversification within the HisB subfamily to identify loop residues that serve as substrate recognition elements and that distinguish GmhB from its subfamily counterpart, the histdinol-phosphate phosphatase domain of HisB. We show that GmhB and the histidinol-phosphate phosphatase domain use the same design of three substrate-recognition loops inserted into the cap domain, yet through selective residue usage on the loops, have achieved unique substrate specificity and thus novel biochemical function.
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影响因子：
2.9
作者：
Dai, Jianying;Finci, Lorenzo;Zhang, Chunchun;Lahiri, Sushmita;Zhang, Guofeng;Peisach, Ezra;Allen, Karen N.;Dunaway-Mariano, Debra
通讯作者：
Dunaway-Mariano, Debra
影响因子：
2.9
作者：
Parsons, JF;Lim, K;Herzberg, O
通讯作者：
Herzberg, O
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：
56.9
作者：
Heine, A;DeSantis, G;Wilson, IA
通讯作者：
Wilson, IA
影响因子：
3.9
作者：
Brilli, M;Fani, R
通讯作者：
Fani, R