Lammens Willem, Le Roy Katrien, Van Laere André, Rabijns Anja, Van den Ende Wim
Laboratorium voor Moleculaire Plantenfysiologie, Faculteit Wetenschappen, Departement Biologie, K.U. Leuven, Kasteelpark Arenberg 31, bus 2434, B-3001 Heverlee, Belgium.
J Mol Biol. 2008 Mar 21;377(2):378-85. doi: 10.1016/j.jmb.2007.12.074. Epub 2008 Jan 5.
In plants, cell-wall invertases fulfil important roles in carbohydrate partitioning, growth, development and crop yield. In this study, we report on different X-ray crystal structures of Arabidopsis thaliana cell-wall invertase 1 (AtcwINV1) mutants with sucrose. These structures reveal a detailed view of sucrose binding in the active site of the wild-type AtcwINV1. Compared to related enzyme-sucrose complexes, important differences in the orientation of the glucose subunit could be observed. The structure of the E203Q AtcwINV1 mutant showed a complete new binding modus, whereas the D23A, E203A and D239A structures most likely represent the productive binding modus. Together with a hydrophobic zone formed by the conserved W20, W47 and W82, the residues N22, D23, R148, E203, D149 and D239 are necessary to create the ideal sucrose-binding pocket. D239 can interact directly with the glucose moiety of sucrose, whereas K242 has an indirect role in substrate stabilization. Most probably, K242 keeps D239 in a favourable position upon substrate binding. Unravelling the exact position of sucrose in plant cell-wall invertases is a necessary step towards the rational design of superior invertases to further increase crop yield and biomass production.
在植物中，细胞壁转化酶在碳水化合物分配、生长、发育和作物产量方面发挥着重要作用。在本研究中，我们报道了拟南芥细胞壁转化酶1（AtcwINV1）突变体与蔗糖的不同X射线晶体结构。这些结构揭示了野生型AtcwINV1活性位点中蔗糖结合的详细情况。与相关的酶 - 蔗糖复合物相比，可以观察到葡萄糖亚基取向的重要差异。E203Q AtcwINV1突变体的结构显示出一种全新的结合模式，而D23A、E203A和D239A结构很可能代表了有效结合模式。与由保守的W20、W47和W82形成的疏水区域一起，残基N22、D23、R148、E203、D149和D239对于创建理想的蔗糖结合口袋是必需的。D239可以直接与蔗糖的葡萄糖部分相互作用，而K242在底物稳定中起间接作用。很可能，K242在底物结合时使D239处于有利位置。揭示蔗糖在植物细胞壁转化酶中的精确位置是合理设计优质转化酶以进一步提高作物产量和生物量生产的必要步骤。
