Bacterial NDH-1 is a key gut target of metformin mediating its antidiabetic effects

Abstract Metformin is the first-line therapy for type 2 diabetes, yet the mechanisms underlying its glucose-lowering effects remain incompletely understood. Increasing evidence implicates gut microbiota as an important mediator of metformin action, but the bacterial targets of the drug remain poorly defined. Here, we identify bacterial NADH oxidoreductase (NDH-1), the homolog of mitochondrial complex I, as a key target of metformin. Metformin inhibited bacterial growth at millimolar concentrations and, at subinhibitory concentrations, markedly impaired carbohydrate fermentation in Escherichia coli and other NDH-1-containing bacteria. Supplementation with nicotinic acid and Mg²⁺ restored fermentation, whereas pyruvate bypassed the metabolic defect, implicating impaired NAD⁺ regeneration and disruption of metabolism upstream of pyruvate formation. Genetic analysis further established NDH-1 dependence: metformin failed to inhibit fermentation in an E. coli ΔnuoF mutant lacking functional NDH-1 but retained activity in an NDH-2-deficient Δndh strain. Molecular docking predicted metformin binding within the NDH-1 quinone-binding region, proximal to conserved residues involved in quinone reduction. Consistent with this mechanism, Staphylococcus aureus , which lacks NDH-1, was not affected in carbohydrate fermentation, while several gut-associated probiotic and short-chain fatty acid-producing bacteria, including Lactobacillus , Bifidobacterium , and Akkermansia muciniphila , lacked detectable NDH-1 homologs, suggesting intrinsic resistance to this metabolic effect of metformin. These findings identify bacterial NDH-1 as a metabolic target of metformin and provide a mechanistic framework for its effects on the gut microbiota. Selective inhibition of NDH-1-dependent bacteria could alter microbial substrate utilization and ecological interactions in the intestine, potentially contributing to the microbiome-mediated component of metformin’s antidiabetic effects. Significance Statement Metformin is one of the most widely used drugs for type 2 diabetes, but how it interacts with the gut microbiota remains unclear. We show that metformin targets bacterial NADH oxidoreductase (NDH-1), the bacterial homolog of mitochondrial complex I, disrupting bacterial energy metabolism and carbohydrate fermentation. Bacteria lacking NDH-1, including gut-associated short-chain fatty acid-producing bacteria such as Lactobacillus , Bifidobacterium , and Akkermansia muciniphila , may therefore be less susceptible to this effect. Our findings identify a bacterial molecular target of metformin and provide a mechanistic framework linking drug-microbe interactions to microbiome remodeling. This work expands the understanding of metformin beyond host-directed mechanisms and highlights bacterial respiratory metabolism as a determinant of drug-microbiota interactions.