Miallau Linda, Hunter William N, McSweeney Sean M, Leonard Gordon A
Macromolecular Crystallography Group, European Synchrotron Radiation Facility, 38043 Grenoble, France.
J Biol Chem. 2007 Jul 6;282(27):19948-57. doi: 10.1074/jbc.M701480200. Epub 2007 Apr 25.
High resolution structures of Staphylococcus aureus d-tagatose-6-phosphate kinase (LacC) in two crystal forms are herein reported. The structures define LacC in apoform, in binary complexes with ADP or the co-factor analogue AMP-PNP, and in a ternary complex with AMP-PNP and D-tagatose-6-phosphate. The tertiary structure of the LacC monomer, which is closely related to other members of the pfkB subfamily of carbohydrate kinases, is composed of a large alpha/beta core domain and a smaller, largely beta "lid." Four extended polypeptide segments connect these two domains. Dimerization of LacC occurs via interactions between lid domains, which come together to form a beta-clasp structure. Residues from both subunits contribute to substrate binding. LacC adopts a closed structure required for phosphoryl transfer only when both substrate and co-factor are bound. A reaction mechanism similar to that used by other phosphoryl transferases is proposed, although unusually, when both substrate and co-factor are bound to the enzyme two Mg(2+) ions are observed in the active site. A new motif of amino acid sequence conservation common to the pfkB subfamily of carbohydrate kinases is identified.
本文报道了金黄色葡萄球菌D-塔格糖-6-磷酸激酶（LacC）两种晶体形式的高分辨率结构。这些结构确定了无配体形式的LacC、与ADP或辅因子类似物AMP-PNP形成的二元复合物中的LacC，以及与AMP-PNP和D-塔格糖-6-磷酸形成的三元复合物中的LacC。LacC单体的三级结构与碳水化合物激酶pfkB亚家族的其他成员密切相关，由一个大的α/β核心结构域和一个较小的、主要为β折叠的“盖子”组成。四个延伸的多肽片段连接这两个结构域。LacC通过盖子结构域之间的相互作用发生二聚化，这些结构域聚集在一起形成一个β扣环结构。两个亚基的残基都参与底物结合。只有当底物和辅因子都结合时，LacC才会采用磷酸转移所需的闭合结构。尽管不同寻常的是，当底物和辅因子都与酶结合时，在活性位点观察到两个Mg(2+)离子，但仍提出了一种与其他磷酸转移酶类似的反应机制。鉴定出了碳水化合物激酶pfkB亚家族共有的一个新的氨基酸序列保守基序。
