STRUCTURAL-ANALYSIS OF THE ZINC HYDROXIDE-THR-199-GLU-106 HYDROGEN-BOND NETWORK IN HUMAN CARBONIC ANHYDRASE-II
STRUCTURAL-ANALYSIS OF THE ZINC HYDROXIDE-THR-199-GLU-106 HYDROGEN-BOND NETWORK IN HUMAN CARBONIC ANHYDRASE-II
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DOI：
10.1002/prot.340170112
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发表时间：
1993-09-01
期刊：
影响因子：
--
通讯作者：
LINDSKOG, S
中科院分区：
文献类型：
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作者：
XUE, YF;LILJAS, A;LINDSKOG, S
通过对位点特异性突变体的X射线晶体学分析，研究了人碳酸酐酶II活性位点中氢氧化锌-Thr-199-Glu-106氢键网络的意义。具有Ala-199和Ala-106或Gln-106的突变体具有低催化活性，而具有Asp-106的突变体具有几乎完全的CO2水合活性。这四个突变体的结构，以及突变体与Ala-199的碳酸氢盐复合物的结构，已被确定在1.7至2.2埃的分辨率。去除残基199的γ原子导致锌离子处的扭曲四面体几何形状，并且催化重要的锌结合水分子已经向Glu-106移动。在突变体与Ala-199的碳酸氢盐复合物中，来自碳酸氢盐的一个氧原子与锌结合，而不取代该水分子。四面体配位几何结构保留在突变体的位置106。具有Ala-106和Gln-106的突变体具有锌结合的硫酸根离子，而该硫酸根位点在具有Asp-166的突变体中仅部分被占据。在Ala-106和Gln-106的突变体中，氢键网络似乎是“颠倒的"。在具有Asp-106的突变体中，该网络与天然酶一样被保留，但是在天然酶中，Asp-106的侧链比Glu-106的侧链延伸得更多。这些结果说明Glu-106和Thr-199对于控制锌离子的精确配位几何形状及其配体偏好的重要性，这导致锌结合的氢氧根离子攻击CO2底物的最佳取向。 (C)1993 Wiley-Liss，Inc.
The significance of the zinc hydroxide-Thr-199-Glu-106 hydrogen-bond network in the active site of human carbonic anhydrase II has been examined by X-ray crystallographic analyses of site-specific mutants. Mutants with Ala-199 and Ala-106 or Gln-106 have low catalytic activities, while a mutant with Asp-106 has almost full CO2 hydration activity. The structures of these four mutants, as well as that of the bicarbonate complex of the mutant with Ala-199, have been determined at 1.7 to 2.2 angstrom resolution. Removal of the gamma atoms of residue 199 leads to a distorted tetrahedral geometry at the zinc ion, and a catalytically important zinc-bound water molecule has moved towards Glu-106. In the bicarbonate complex of the mutant with Ala-199 one oxygen atom from bicarbonate binds to zinc without displacing this water molecule. Tetrahedral coordination geometries are retained in the mutants at position 106. The mutants with Ala-106 and Gln-106 have a zinc-bound sulfate ion, whereas this sulfate site is only partially occupied in the mutant with Asp-166. The hydrogen-bond network seems to be ''reversed'' in the mutants with Ala-106 and Gln-106. The network is preserved as in native enzyme in the mutant with Asp-106 but the side chain of Asp-106 is more extended than that of Glu-106 in the native enzyme. These results illustrate the importance of Glu-106 and Thr-199 for controlling the precise coordination geometry of the zinc ion and its ligand preferences which results in an optimal orientation of a zinc-bound hydroxide ion for an attack on the CO2 substrate. (C) 1993 Wiley-Liss, Inc.