Verhaest Maureen, Lammens Willem, Le Roy Katrien, De Coninck Barbara, De Ranter Camiel J, Van Laere André, Van den Ende Wim, Rabijns Anja
Laboratorium voor Biokristallografie, Faculteit Farmaceutische Wetenschappen, K. U. Leuven, Herestraat 49, O&N II, Bus 822, B-3000 Leuven, Belgium.
Acta Crystallogr D Biol Crystallogr. 2006 Dec;62(Pt 12):1555-63. doi: 10.1107/S0907444906044489. Epub 2006 Nov 23.
Cell-wall invertases play crucial roles during plant development. They hydrolyse sucrose into its fructose and glucose subunits by cleavage of the alpha1-beta2 glycosidic bond. Here, the structure of the Arabidopsis thaliana cell-wall invertase 1 (AtcwINV1; gene accession code At3g13790) is described at a resolution of 2.15 A. The structure comprises an N-terminal fivefold beta-propeller domain followed by a C-terminal domain formed by two beta-sheets. The active site is positioned in the fivefold beta-propeller domain, containing the nucleophile Asp23 and the acid/base catalyst Glu203 of the double-displacement enzymatic reaction. The function of the C-terminal domain remains unknown. Unlike in other GH 32 family enzyme structures known to date, in AtcwINV1 the cleft formed between both domains is blocked by Asn299-linked carbohydrates. A preliminary site-directed mutagenesis experiment (Asn299Asp) removed the glycosyl chain but did not alter the activity profile of the enzyme.
细胞壁转化酶在植物发育过程中发挥着关键作用。它们通过切割α1-β2糖苷键将蔗糖水解为果糖和葡萄糖亚基。在此,以2.15 Å的分辨率描述了拟南芥细胞壁转化酶1(AtcwINV1;基因登录号At3g13790)的结构。该结构包括一个N端五倍体β-螺旋桨结构域,其后是由两个β-折叠形成的C端结构域。活性位点位于五倍体β-螺旋桨结构域中,包含双置换酶促反应的亲核试剂Asp23和酸碱催化剂Glu203。C端结构域的功能仍然未知。与迄今为止已知的其他GH 32家族酶结构不同,在AtcwINV1中,两个结构域之间形成的裂隙被Asn299连接的碳水化合物封闭。初步的定点诱变实验(Asn299Asp)去除了糖基链,但没有改变该酶的活性谱。