Chumanevich Alexander A, Krupenko Sergey A, Davies Christopher
Department of Biochemistry and Molecular Biology, Medical University of South Carolina, Charleston, South Carolina 29425, USA.
J Biol Chem. 2004 Apr 2;279(14):14355-64. doi: 10.1074/jbc.M313934200. Epub 2004 Jan 16.
10-Formyltetrahydrofolate dehydrogenase (FDH) converts 10-formyltetrahydrofolate, a precursor for nucleotide biosynthesis, to tetrahydrofolate. The protein comprises two functional domains: a hydrolase domain that removes a formyl group from 10-formyltetrahydrofolate and a NADP(+)-dependent dehydrogenase domain that reduces the formyl to carbon dioxide. As a first step toward deciphering the catalytic mechanism of the enzyme, we have determined the crystal structure of the hydrolase domain of FDH from rat, solved to 2.3-A resolution. The structure comprises two domains. As expected, domain 1 shares the same Rossmann fold as the related enzymes, methionyl-tRNA-formyltransferase and glycinamide ribonucleotide formyltransferase, but, unexpectedly, the structural similarity between the amino-terminal domain of 10-formyltetrahydrofolate dehydrogenase and methionyl-tRNA-formyltransferase extends to the C terminus of both proteins. The active site contains a molecule of beta-mercaptoethanol that is positioned between His-106 and Asp-142 and that appears to mimic the formate product. We propose a catalytic mechanism for the hydrolase reaction in which Asp-142 polarizes the catalytic water molecule and His-106 orients the carbonyl group of formyl. The structure also provides clues as to how, in the native enzyme, the hydrolase domain transfers its product to the dehydrogenase domain.
10-甲酰四氢叶酸脱氢酶（FDH）将核苷酸生物合成的前体10-甲酰四氢叶酸转化为四氢叶酸。该蛋白质包含两个功能结构域：一个从10-甲酰四氢叶酸去除甲酰基的水解酶结构域和一个将甲酰基还原为二氧化碳的依赖NADP(+)的脱氢酶结构域。作为解析该酶催化机制的第一步，我们确定了大鼠FDH水解酶结构域的晶体结构，分辨率达到2.3埃。该结构包含两个结构域。正如预期的那样，结构域1与相关酶甲硫氨酰-tRNA-甲酰基转移酶和甘氨酰胺核糖核苷酸甲酰基转移酶具有相同的Rossmann折叠，但出乎意料的是，10-甲酰四氢叶酸脱氢酶的氨基末端结构域与甲硫氨酰-tRNA-甲酰基转移酶之间的结构相似性延伸到了这两种蛋白质的C末端。活性位点含有一个β-巯基乙醇分子，它位于His-106和Asp-142之间，似乎模拟了甲酸产物。我们提出了一种水解酶反应的催化机制，其中Asp-142使催化水分子极化，His-106使甲酰基的羰基定向。该结构还提供了关于在天然酶中水解酶结构域如何将其产物转移到脱氢酶结构域的线索。
