The molecular architecture of human N-acetylgalactosamine kinase
The molecular architecture of human N-acetylgalactosamine kinase
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DOI：
10.1074/jbc.m505730200
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发表时间：
2005-09-23
影响因子：
4.8
通讯作者：
Holden, HM
中科院分区：
文献类型：
--
作者：
Thoden, JB;Holden, HM
半乳糖激酶通过催化 α-(D)-半乳糖转化为半乳糖 1-磷酸，在正常半乳糖代谢中发挥关键作用。近年来，人类半乳糖激酶和该酶的两种细菌形式的三维结构已被确定。最初，编码人类半乳糖激酶的基因被定位在 17 号染色体上。随后，另一个编码与半乳糖激酶序列相似的蛋白质的基因被定位在 15 号染色体上。最近的报告表明，第二个基因 (GALK2) 编码的酶对 GalNAc 的活性比对半乳糖的活性更强。这种酶（GalNAc 激酶）与复杂碳水化合物降解产生的游离 GalNAc 的再利用的挽救途径有关。在这里，我们报告了 GalNAc 激酶的首次结构分析。人类酶的结构在 MnAMPPNP 和 GalNAc 或 MgATP 和 GalNAc 存在的情况下得到解析（这会在活性位点产生结合产物）。该酶表现出明显的双叶外观，其活性位点楔入两个结构域之间。 N 端区域以七链混合 β 片层为主，而 C 端基序包含两层反平行 β 片层。 GalNAc 激酶显示的整体拓扑结构将其置于酶的 GHMP 超家族中，该超家族通常起到小分子激酶的作用。通过这项研究，揭示了 GalNAc 激酶活性位点在催化前后的几何形状，并在分子水平上定义了底物特异性的决定因素。
Galactokinase plays a key role in normal galactose metabolism by catalyzing the conversion of alpha-(D)-galactose to galactose 1-phosphate. Within recent years, the three-dimensional structures of human galactokinase and two bacterial forms of the enzyme have been determined. Originally, the gene encoding galactokinase in humans was mapped to chromosome 17. An additional gene, encoding a protein with sequence similarity to galactokinase, was subsequently mapped to chromosome 15. Recent reports have shown that this second gene (GALK2) encodes an enzyme with greater activity against GalNAc than galactose. This enzyme, GalNAc kinase, has been implicated in a salvage pathway for the reutilization of free GalNAc derived from the degradation of complex carbohydrates. Here we report the first structural analysis of a GalNAc kinase. The structure of the human enzyme was solved in the presence of MnAMPPNP and GalNAc or MgATP and GalNAc ( which resulted in bound products in the active site). The enzyme displays a distinctly bilobal appearance with its active site wedged between the two domains. The N- terminal region is dominated by a seven-stranded mixed beta-sheet, whereas the C-terminal motif contains two layers of anti-parallel beta-sheet. The overall topology displayed by GalNAc kinase places it into the GHMP superfamily of enzymes, which generally function as small molecule kinases. From this investigation, the geometry of the GalNAc kinase active site before and after catalysis has been revealed, and the determinants of substrate specificity have been defined on a molecular level.