Dual-Target Inhibitors of the Folate Pathway Inhibit Intrinsically Trimethoprim-Resistant DfrB Dihydrofolate Reductases

Abstract

Trimethoprim (TMP) is widely used to treat infections in humans and in livestock, accelerating the incidence of TMP resistance. The emergent and largely untracked type II dihydrofolate reductases (DfrBs) are intrinsically TMP-resistant plasmid-borne Dfrs that are structurally and evolutionarily unrelated to chromosomal Dfrs. We report kinetic characterization of the known DfrB family members. Their kinetic constants are conserved and all are poorly inhibited by TMP, consistent with TMP resistance. We investigate their inhibition with known and novel bisubstrate inhibitors of 6-hydroxymethyl-7,8-dihydropterin pyrophosphokinase (HPPK). Importantly, all are inhibited by the HPPK inhibitors, making these molecules dual-target inhibitors of two folate pathway enzymes that are strictly microbial.

A dual-target herbicidal inhibitor of lysine biosynthesis

“…Furthermore, the low-micromolar potency of MBDTA-2 at the DHDPR target site is comparable to the potency of glyphosate, the most successful commercial herbicide active ingredient, at its enzyme target 5-enolpyruvylshikimate-3-phosphate synthase ( Mao et al, 2016 ; Sammons et al, 2018 ). The phenomenon of inhibitors having dual-target activity against consecutive enzymes in metabolic pathways has sometimes been attributed to conserved active site features ( Hsu et al, 2013 ; Toulouse et al, 2020 ). However, there are also many examples of inhibitors of multiple targets from distinct pathways, which have been identified regardless of binding site similarities ( Allen et al, 2015 ; Hashmi et al, 2021 ; Wang et al, 2019 ).…”

A dual-target herbicidal inhibitor of lysine biosynthesis

“…Moreover, the ~8-fold greater potency of MBDTA-2 against DHDPR than DHDPS reveals that the in vitro potency is actually ~6-fold greater than the in vivo potency. The phenomenon of inhibitors having dual-target activity against consecutive enzymes in metabolic pathways has sometimes been attributed to conserved active site features (Hsu et al, 2013; Toulouse et al, 2020). However, there are also many examples of inhibitors of multiple targets from distinct pathways, which have been identified regardless of binding site similarities (Allen et al, 2015; Hashmi et al, 2021; Wang et al, 2019).…”

Hfq Regulates Efflux Pump Expression and Purine Metabolic Pathway to Increase Trimethoprim Resistance in Aeromonas veronii

“…But in fact, the productions of purines and nucleotides are affected by one carbon unit carrier tetrahydrofolate, and the latter is controlled by dihydrofolate reductase in turn ( Paulsen et al, 2013 ). Since antibiotic trimethoprim targets dihydrofolate reductase specifically ( Darrell et al, 1968 ; Bourne, 2014 ; Toulouse et al, 2020 ), trimethoprim is treated for Hfq knockout instead of other antibiotics. There are many mechanisms for resistance, of which efflux pump is important for multidrug resistance in bacteria ( Elsby et al, 2017 ).…”