Crystal structures of complexes with cobalt-reconstituted human arginase I.
Crystal structures of complexes with cobalt-reconstituted human arginase I.
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DOI：
10.1021/bi201101t
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发表时间：
2011-09-20
期刊：
影响因子：
2.9
通讯作者：
Christianson, David W.
中科院分区：
文献类型：
--
作者：
D'Antonio, Edward L.;Christianson, David W.
双核锰金属酶人精氨酸酶I （HAI）是一种潜在的癌症化疗蛋白药物，因为它能够消耗肿瘤细胞微环境中需要这种营养物质才能茁壮成长的细胞外l-精氨酸水平。在活性位点用Co2+2簇取代天然Mn2+2簇产生一种酶，在生理pH（~7.4）下具有增强的催化活性，可以作为l-精氨酸耗损治疗的改进蛋白药物。与Mn2+2-HAI相比，Co2+2-HAI的催化机理不同，包括不同寻常的Nε—Co2+配位模式，以解释l-Arg较低的KM值和l-Orn较低的Ki值。然而，我们现在报道在钴重组酶中没有观察到异常的金属配位模式：在2.10 Å分辨率（pH 7.0）和1.97 Å分辨率（pH 8.5）下测定的无配体Co2+2- hai的x射线晶体结构，以及在1.85 Å和1.50 Å分辨率下测定的与反应底物类似物2(S)-氨基-6-硼已酸（ABH, pH 7.0）和催化产物l-Orn （pH 7.0）配合的结构，与Mn2+2- hai的相应结构基本相同。因此，在Co2+2-HAI和Mn2+2-HAI之间没有明显的结构差异的情况下，我们认为Co2+2-HAI在生理pH下具有更高浓度的金属桥接氢氧化物离子（这是Co2+结合水分子比Mn2+结合水分子具有更低的pKa的结果）加强了与阳离子氨基酸的静电相互作用，并导致亲和力增强，反映在l-Arg的较低KM值和l-Orn的较低Ki值上。
The binuclear manganese metalloenzyme human arginase I (HAI) is a potential protein drug for cancer chemotherapy, in that it is capable of depleting extracellular l-Arg levels in the microenvironment of tumor cells that require this nutrient to thrive. Substitution of the native Mn2+2 cluster with a Co2+2 cluster in the active site yields an enzyme with enhanced catalytic activity at physiological pH (~7.4) that could serve as an improved protein drug for l-Arg depletion therapy. A different catalytic mechanism is proposed for Co2+2-HAI compared with that of Mn2+2-HAI, including an unusual Nε---Co2+ coordination mode, to rationalize the lower KM value of l-Arg and the lower Ki value of l-Orn. However, we now report that no unusual metal coordination modes are observed in the cobalt-reconstituted enzyme: the X-ray crystal structures of unliganded Co2+2-HAI determined at 2.10 Å resolution (pH 7.0) and 1.97 Å resolution (pH 8.5), as well as the structures of Co2+2-HAI complexed with the reactive substrate analogue 2(S)-amino-6-boronohexanoic acid (ABH, pH 7.0) and the catalytic product l-Orn (pH 7.0) determined at 1.85 Å resolution and 1.50 Å resolution, respectively, are essentially identical to the corresponding structures of Mn2+2-HAI. Therefore, in the absence of significant structural differences between Co2+2-HAI and Mn2+2-HAI, we suggest that a higher concentration of metal-bridging hydroxide ion at physiological pH for Co2+2-HAI – a consequence of the lower pKa of a Co2+-bound water molecule compared with a Mn2+-bound water molecule – strengthens electrostatic interactions with cationic amino acids and accounts for enhanced affinity as reflected in the lower KM value of l-Arg and the lower Ki value of l-Orn.
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影响因子：
7.3
作者：
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影响因子：
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影响因子：
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DOI：
10.1107/s0907444904019158
发表时间：
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影响因子：
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作者：
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