Sigrell J A, Cameron A D, Jones T A, Mowbray S L
Department of Molecular Biology, Uppsala University, Sweden.
Structure. 1998 Feb 15;6(2):183-93. doi: 10.1016/s0969-2126(98)00020-3.
D-ribose must be phosphorylated at O5' before it can be used in either anabolism or catabolism. This reaction is catalysed by ribokinase and requires the presence of ATP and magnesium. Ribokinase is a member of a family of carbohydrate kinases of previously unknown structure.
The crystal structure of ribokinase from Escherichia coli in complex with ribose and dinucleotide was determined at 1.84 A resolution by multiple isomorphous replacement. There is one 33 kDa monomer of ribokinase in the asymmetric unit but the protein forms a dimer around a crystallographic twofold axis. Each subunit consists of a central alpha/beta unit, with a new type of nucleotide-binding fold, and a distinct beta sheet that forms a lid over the ribose-binding site. Contact between subunits involves orthogonal packing of beta sheets, in a novel dimer interaction that we call a beta clasp.
Inspection of the complex indicates that ribokinase utilises both a catalytic base for activation of the ribose in nucleophilic attack and an anion hole that stabilises the transition state during phosphoryl transfer. The structure suggests an ordered reaction mechanism, similar to those proposed for other carbohydrate kinases that probably involves conformational changes. We propose that the beta-clasp structure acts as a lid, closing and opening upon binding and release of ribose. From these observations, an understanding of the structure and catalytic mechanism of related sugar kinases can be obtained.
D - 核糖在用于合成代谢或分解代谢之前必须在O5'位被磷酸化。此反应由核糖激酶催化，需要ATP和镁的存在。核糖激酶是一个结构此前未知的碳水化合物激酶家族的成员。
通过多同晶置换法，以1.84 Å的分辨率测定了来自大肠杆菌的核糖激酶与核糖和二核苷酸复合物的晶体结构。不对称单元中有一个33 kDa的核糖激酶单体，但该蛋白围绕一个晶体学二重轴形成二聚体。每个亚基由一个中央α/β单元和一个独特的β折叠片组成，中央α/β单元具有一种新型的核苷酸结合折叠，β折叠片在核糖结合位点上方形成一个盖子。亚基之间的接触涉及β折叠片的正交堆积，形成一种我们称为β扣的新型二聚体相互作用。
对该复合物的研究表明，核糖激酶在亲核攻击中利用一个催化碱基来激活核糖，并利用一个阴离子空穴在磷酸转移过程中稳定过渡态。该结构表明了一种有序的反应机制，类似于为其他碳水化合物激酶所提出的机制，可能涉及构象变化。我们提出β扣结构起到盖子的作用，在核糖结合和释放时开合。基于这些观察结果，可以了解相关糖激酶的结构和催化机制。
