Université de Strasbourg, CNRS UMR 7177, Institut Le Bel, 4 rue Blaise Pascal, CS 90032, 67081 Strasbourg Cedex, France.
J Am Chem Soc. 2013 Feb 6;135(5):1816-22. doi: 10.1021/ja309557s. Epub 2013 Jan 29.
The MEP pathway, which is absent in animals but present in most pathogenic bacteria, in the parasite responsible for malaria and in plant plastids, is a target for the development of antimicrobial drugs. IspH, an oxygen-sensitive [4Fe-4S] enzyme, catalyzes the last step of this pathway and converts (E)-4-hydroxy-3-methylbut-2-en-1-yl diphosphate (HMBPP) into the two isoprenoid precursors: isopentenyl diphosphate (IPP) and dimethylallyl diphosphate (DMAPP). A crucial step in the mechanism of this enzyme is the binding of the C4 hydroxyl of HMBPP to the unique fourth iron site in the 4Fe-4S moiety. Here, we report the synthesis and the kinetic investigations of two new extremely potent inhibitors of E. coli IspH where the OH group of HMBPP is replaced by an amino and a thiol group. (E)-4-Mercapto-3-methylbut-2-en-1-yl diphosphate is a reversible tight-binding inhibitor of IspH with K(i) = 20 ± 2 nM. A detailed kinetic analysis revealed that (E)-4-amino-3-methylbut-2-en-1-yl diphosphate is a reversible slow-binding inhibitor of IspH with K(i) = 54 ± 19 nM. The slow binding behavior of this inhibitor is best described by a one-step mechanism with the slow step consisting of the formation of the enzyme-inhibitor (EI) complex.
MEP 途径在动物中不存在，但在大多数致病细菌、疟原虫和植物质体中存在，是开发抗菌药物的靶点。IspH 是一种对氧气敏感的[4Fe-4S]酶，催化该途径的最后一步，将（E）-4-羟基-3-甲基-2-丁烯-1-基二磷酸（HMBPP）转化为两种异戊烯基前体：异戊烯二磷酸（IPP）和二甲基烯丙基二磷酸（DMAPP）。该酶的机制中一个关键步骤是 HMBPP 的 C4 羟基与 4Fe-4S 部分中独特的第四个铁位结合。在这里，我们报告了两种新型强效大肠杆菌 IspH 抑制剂的合成和动力学研究，其中 HMBPP 的 OH 基团被氨基和巯基取代。（E）-4-巯基-3-甲基-2-丁烯-1-基二磷酸是 IspH 的可逆紧密结合抑制剂，K(i) = 20 ± 2 nM。详细的动力学分析表明，（E）-4-氨基-3-甲基-2-丁烯-1-基二磷酸是 IspH 的可逆慢结合抑制剂，K(i) = 54 ± 19 nM。该抑制剂的慢结合行为最好用一步机制来描述，其中慢步骤包括酶-抑制剂（EI）复合物的形成。
