Chen FangFang, Wang Ping, An Yan, Huang JianQin, Xu YingWu
The Nurturing Station for the State Key Laboratory of Subtropical Sylviculture, Zhejiang Agriculture and Forestry University, Lin'an, Zhejiang 311300, China.
The Nurturing Station for the State Key Laboratory of Subtropical Sylviculture, Zhejiang Agriculture and Forestry University, Lin'an, Zhejiang 311300, China.
Biochimie. 2015 Jan;108:33-9. doi: 10.1016/j.biochi.2014.10.023. Epub 2014 Nov 4.
Alcohol dehydrogenase (ADH, EC 1.1.1.1) plays important roles in the metabolism of alcohols and aldehydes. They are often subjected to conformational changes that are critical for the enzymatic activity and have received intensive investigation for horse liver ADH. However, for the large plant ADH members, little is known regarding both the conformational change and its relationship to catalytic activity as plant ADH structures were rarely available. Here we describe three Arabidopsis ADH conformations obtained from two crystals, the apo crystal that was free of ligand, and the complex crystal that was with NAD. The NAD-complexed crystal yielded two different structural forms for the two subunits, one was occupied by the coenzyme, and the other was free and open. Structural comparisons demonstrate that the occupied subunit is in a closed conformation while the free subunit is fully open, and the apo structure in intermediate. Though all the forms have an overall fold similar to that of horse and human ADHs, the catalytic domain has an over 10° rotation. Additionally, unlike horse liver ADH, the loop (295-302aa) adopts different conformation. It does not rearrange upon the binding of the coenzyme norVal297 side chain experiences a flipping. Instead it always remains in the active site. His48 plays a switching role in the structure. Its imidazole ring has to swim away from the binding site to permit NAD binding. These together with the large differences in the substrate binding pocket, as well as in the proton relay system demonstrate that AtADH adopts a different catalysis mechanism from horse liver ADH.
乙醇脱氢酶（ADH，EC 1.1.1.1）在醇类和醛类的代谢中发挥着重要作用。它们经常经历构象变化，这对酶活性至关重要，并且马肝ADH已受到深入研究。然而，对于大型植物ADH成员，由于很少有植物ADH结构可用，关于构象变化及其与催化活性的关系知之甚少。在这里，我们描述了从两种晶体中获得的三种拟南芥ADH构象，一种是不含配体的脱辅基晶体，另一种是与NAD结合的复合晶体。NAD复合晶体为两个亚基产生了两种不同的结构形式，一种被辅酶占据，另一种是游离且开放的。结构比较表明，被占据的亚基处于封闭构象，而游离亚基完全开放，脱辅基结构处于中间状态。尽管所有形式的总体折叠与马和人ADH相似，但催化结构域有超过10°的旋转。此外，与马肝ADH不同，环（295 - 302aa）采用不同的构象。它在辅酶结合时不会重新排列，也没有观察到Val297侧链的翻转。相反，它始终保留在活性位点。His48在结构中起转换作用。其咪唑环必须从结合位点移开以允许NAD结合。这些与底物结合口袋以及质子传递系统中的巨大差异一起表明，AtADH采用了与马肝ADH不同的催化机制。
