Vinnicombe H G, Derrick J P
Department of Biomolecular Sciences, UMIST, Manchester, M60 1QW, United Kingdom.
Biochem Biophys Res Commun. 1999 May 19;258(3):752-7. doi: 10.1006/bbrc.1999.0695.
Dihydropteroate synthase (DHPS) catalyses a key step in the biosynthesis of folic acid and is the target for inhibition by the sulphonamide class of antimicrobial agents. Here we describe a study of the enzymatic mechanism and sulphonamide inhibition of DHPS from the pathogen Streptococcus pneumoniae. Equilibrium binding assays showed that binding of the substrate para-aminobenzoic acid (pABA) to DHPS was absolutely dependent on the presence of pyrophosphate, which acts as an analogue of the second substrate 6-hydroxymethyl-7, 8-dihydropterin pyrophosphate (DHPPP). The product of the reaction, dihydropteroate, was also able to bind to DHPS. Sulphonamides were capable of displacing pABA in a competitive manner, with equilibrium binding constants that were significantly higher than the equivalent Ki values deduced from steady state kinetic measurements. These results indicate that the target for sulphonamide inhibition of S. pneumoniae DHPS is the enzyme-DHPPP binary complex, rather than the apoprotein form of the enzyme.
二氢蝶酸合酶（DHPS）催化叶酸生物合成中的关键步骤，是磺胺类抗菌剂的抑制靶点。在此，我们描述了一项关于病原菌肺炎链球菌DHPS的酶促机制及磺胺抑制作用的研究。平衡结合试验表明，底物对氨基苯甲酸（pABA）与DHPS的结合绝对依赖于焦磷酸的存在，焦磷酸作为第二种底物6-羟甲基-7,8-二氢蝶呤焦磷酸（DHPPP）的类似物。反应产物二氢蝶酸也能够与DHPS结合。磺胺类药物能够以竞争性方式取代pABA，其平衡结合常数显著高于从稳态动力学测量推导得出的等效Ki值。这些结果表明，磺胺类药物对肺炎链球菌DHPS的抑制靶点是酶-DHPPP二元复合物，而非酶的脱辅基蛋白形式。
