7KL8

Structure of F420 binding protein Rv1558 from Mycobacterium tuberculosis with F420 bound

Summary for 7KL8

• Entry DOI: 10.2210/pdb7kl8/pdb
• Descriptor: Deazaflavin-dependent nitroreductase, COENZYME F420-3, SULFATE ION, ... (5 entities in total)
• Functional Keywords: f420, fdor, oxidoreductase
• Biological source: Mycobacterium tuberculosis
• Total number of polymer chains: 2
• Total formula weight: 33644.05

Authors

Lee, B.M.,Tan, L.L.,Jackson, C.J. (deposition date: 2020-10-29, release date: 2021-03-24, Last modification date: 2023-10-18)

Primary citation

Aragaw, W.W.,Lee, B.M.,Yang, X.,Zimmerman, M.D.,Gengenbacher, M.,Dartois, V.,Chui, W.K.,Jackson, C.J.,Dick, T.
Potency boost of a Mycobacterium tuberculosis dihydrofolate reductase inhibitor by multienzyme F 420 H 2 -dependent reduction.
Proc.Natl.Acad.Sci.USA, 118:-, 2021

PubMed Abstract: Triaza-coumarin (TA-C) is a (Mtb) dihydrofolate reductase (DHFR) inhibitor with an IC (half maximal inhibitory concentration) of ∼1 µM against the enzyme. Despite this moderate target inhibition, TA-C shows exquisite antimycobacterial activity (MIC, concentration inhibiting growth by 50% = 10 to 20 nM). Here, we investigated the mechanism underlying this potency disconnect. To confirm that TA-C targets DHFR and investigate its unusual potency pattern, we focused on resistance mechanisms. In Mtb, resistance to DHFR inhibitors is frequently associated with mutations in thymidylate synthase , which sensitizes Mtb to DHFR inhibition, rather than in DHFR itself. We observed mutations, consistent with TA-C interfering with the folate pathway. A second resistance mechanism involved biosynthesis of the redox coenzyme F Thus, we hypothesized that TA-C may be metabolized by Mtb F-dependent oxidoreductases (FDORs). By chemically blocking the putative site of FDOR-mediated reduction in TA-C, we reproduced the F-dependent resistance phenotype, suggesting that FH-dependent reduction is required for TA-C to exert its potent antibacterial activity. Indeed, chemically synthesized TA-C-Acid, the putative product of TA-C reduction, displayed a 100-fold lower IC against DHFR. Screening seven recombinant Mtb FDORs revealed that at least two of these enzymes reduce TA-C. This redundancy in activation explains why no mutations in the activating enzymes were identified in the resistance screen. Analysis of the reaction products confirmed that FDORs reduce TA-C at the predicted site, yielding TA-C-Acid. This work demonstrates that intrabacterial metabolism converts TA-C, a moderately active "prodrug," into a 100-fold-more-potent DHFR inhibitor, thus explaining the disconnect between enzymatic and whole-cell activity.

PubMed: 34161270
DOI: 10.1073/pnas.2025172118

Experimental method

X-RAY DIFFRACTION (2.469 Å)