Department of Biochemistry, University of Cambridge, Tennis Court Road, Cambridge CB2 1QA, UK; Laboratório Nacional de Biociências (LNBio), Centro de Nacional de Pesquisa em Energia e Materiais, Campinas, (CNPEM), São Paulo 13083-100, Brazil.
Department of Biochemistry, University of Cambridge, Tennis Court Road, Cambridge CB2 1QA, UK.
Structure. 2014 Jan 7;22(1):94-103. doi: 10.1016/j.str.2013.09.022. Epub 2013 Nov 7.
Inhibition of the biosynthesis of tetrahydrofolate (THF) has long been a focus in the treatment of both cancer and infectious diseases. Dihydrofolate reductase (DHFR), which catalyzes the last step, is one of the most thoroughly explored targets of this pathway, but there are no DHFR inhibitors used for tuberculosis treatment. Here, we report a structural, site-directed mutagenesis and calorimetric analysis of Mycobacterium tuberculosis DHFR (MtDHFR) in complex with classical DHFR inhibitors. Our study provides insights into the weak inhibition of MtDHFR by trimethoprim and other antifolate drugs, such as pyrimethamine and cycloguanil. The construction of the mutant Y100F, together with calorimetric studies, gives insights into low affinity of MtDHFR for classical DHFR inhibitors. Finally, the structures of MtDHFR in complex with pyrimethamine and cycloguanil define important interactions in the active site and provide clues to the more effective design of antibiotics targeted against MtDHFR.
四氢叶酸（THF）的生物合成抑制一直是癌症和传染病治疗的重点。催化最后一步的二氢叶酸还原酶（DHFR）是该途径中研究最透彻的靶点之一，但目前尚无用于结核病治疗的 DHFR 抑制剂。在这里，我们报告了结核分枝杆菌 DHFR（MtDHFR）与经典 DHFR 抑制剂结合的结构、定点突变和量热分析。我们的研究深入了解了甲氧苄啶和其他抗叶酸药物（如乙胺嘧啶和环胍）对 MtDHFR 的抑制作用较弱。构建 Y100F 突变体以及量热研究深入了解了 MtDHFR 对经典 DHFR 抑制剂的低亲和力。最后，MtDHFR 与乙胺嘧啶和环胍复合物的结构定义了活性位点中的重要相互作用，并为更有效地设计针对 MtDHFR 的抗生素提供了线索。
