Valderas Michelle Wright, Andi Babak, Barrow William W, Cook Paul F
Department of Veterinary Pathobiology, Center for Veterinary Health Sciences, Oklahoma State University, 250 Mc Elroy Hall, Stillwater, Oklahoma 74078, USA.
Biochim Biophys Acta. 2008 May;1780(5):848-53. doi: 10.1016/j.bbagen.2008.02.003. Epub 2008 Mar 10.
Dihydropteroate synthase (DHPS) catalyzes the formation of dihydropteroate and Mg-pyrophosphate from 6-hydroxymethyl-7,8-dihydropterin diphosphate and para-aminobenzoic acid. The Bacillus anthracis DHPS is intrinsically resistant to sulfonamides. However, using a radioassay that monitors the dihydropteroate product, the enzyme was inhibited by the same sulfonamides. A continuous spectrophotometric assay for measuring the enzymatic activity of DHPS was developed and used to examine the effects of sulfonamides on the enzyme. The new assay couples the production of MgPPi to the pyrophosphate-dependent phosphofructokinase/aldolase/triose isomerase/alpha-glycerophosphate dehydrogenase reactions and monitors the disappearance of NADH at 340nm. The coupled enzyme assay demonstrates that resistance of the B. anthracis DHPS results in part from the use of the sulfonamides as alternative substrates, resulting in the formation of sulfonamide-pterin adducts, and not necessarily due to an inability to bind them.
二氢蝶酸合酶（DHPS）催化由6-羟甲基-7,8-二氢蝶呤二磷酸和对氨基苯甲酸形成二氢蝶酸和焦磷酸镁。炭疽芽孢杆菌的DHPS对磺胺类药物具有内在抗性。然而，使用监测二氢蝶酸产物的放射性测定法，该酶被相同的磺胺类药物抑制。开发了一种用于测量DHPS酶活性的连续分光光度测定法，并用于研究磺胺类药物对该酶的影响。新的测定法将焦磷酸镁的产生与焦磷酸依赖性磷酸果糖激酶/醛缩酶/磷酸丙糖异构酶/α-甘油磷酸脱氢酶反应偶联，并监测340nm处NADH的消失。偶联酶测定表明，炭疽芽孢杆菌DHPS的抗性部分源于将磺胺类药物用作替代底物，导致形成磺胺类-蝶呤加合物，而不一定是由于无法结合它们。
