Hövel Klaus, Shallom Dalia, Niefind Karsten, Belakhov Valery, Shoham Gil, Baasov Timor, Shoham Yuval, Schomburg Dietmar
Institute for Biochemistry, University of Cologne, Cologne 50674, Germany.
EMBO J. 2003 Oct 1;22(19):4922-32. doi: 10.1093/emboj/cdg494.
High-resolution crystal structures of alpha-L-arabinofuranosidase from Geobacillus stearothermophilus T-6, a family 51 glycosidase, are described. The enzyme is a hexamer, and each monomer is organized into two domains: a (beta/alpha)8-barrel and a 12-stranded beta sandwich with jelly-roll topology. The structures of the Michaelis complexes with natural and synthetic substrates, and of the transient covalent arabinofuranosyl-enzyme intermediate represent two stable states in the double displacement mechanism, and allow thorough examination of the catalytic mechanism. The arabinofuranose sugar is tightly bound and distorted by an extensive network of hydrogen bonds. The two catalytic residues are 4.7 A apart, and together with other conserved residues contribute to the stabilization of the oxocarbenium ion-like transition state via charge delocalization and specific protein-substrate interactions. The enzyme is an anti-protonator, and a 1.7 A electrophilic migration of the anomeric carbon takes place during the hydrolysis.
本文描述了嗜热栖热放线菌T-6来源的α-L-阿拉伯呋喃糖苷酶（一种51家族糖苷酶）的高分辨率晶体结构。该酶是一种六聚体，每个单体由两个结构域组成：一个（β/α）8桶状结构域和一个具有果冻卷拓扑结构的12链β折叠三明治结构域。与天然和合成底物形成的米氏复合物以及瞬态共价阿拉伯呋喃糖基 - 酶中间体的结构代表了双取代机制中的两种稳定状态，并允许对催化机制进行深入研究。阿拉伯呋喃糖通过广泛的氢键网络紧密结合并发生扭曲。两个催化残基相距4.7 Å，与其他保守残基一起通过电荷离域和特定的蛋白质 - 底物相互作用，有助于稳定氧鎓离子样过渡态。该酶是一种反质子供体，在水解过程中异头碳发生1.7 Å的亲电迁移。
