Biology subjects

Pinault, L.

Publications and source records attributed to Pinault, L..

5 recordsLinked to original sources

bioRxiv2026

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

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 delta-nuoF mutant lacking functional NDH-1 but retained activity in an NDH-2-deficient delta-ndh 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.

microbiology↗

bioRxiv2026

Methyltransferases in Candidate Phyla Radiation: A Weapon or a Simple Shield?

The Candidate Phyla Radiation (CPR) consists of microorganisms with highly reduced genomes and limited metabolic capabilities, including the inability to synthesize nucleotides and amino acids. Previous studies revealed an unexpected abundance of antimicrobial resistance-related genes in CPR genomes, with methyltransferases (MTs) accounting for up to 47.34% of detected resistance determinants. Here, we investigated the distribution, diversity, and potential functions of MTs across 12,552 CPR genomes, focusing on their roles in antimicrobial resistance and restriction-modification (R-M) systems. Sequence similarity searches were performed using BLASTp, followed by domain validation with the NCBI Conserved Domain Database (CDD). Putative restriction endonucleases (REs) associated with these systems were also identified and analyzed. We detected 20,232 resistance-associated MTs, 7,391 hsdM-like MTs, and 4,737 REs, revealing a high prevalence of putatively functional MTs relative to REs across CPR genomes. Functional CPR MTs and REs displayed extensive sequence divergence from known bacterial homologs and exhibited greater domain diversity and functional complexity than their bacterial counterparts. Selected high-confidence MT candidates were heterologously expressed in Escherichia coli BL21 and characterized using BIOLOG phenotypic assays and antimicrobial susceptibility testing. Transformed strains showed enhanced utilization of methyl pyruvate as a carbon source, whereas no significant antimicrobial resistance phenotype was detected. Collectively, these findings suggest that CPR-encoded MTs are primarily associated with R-M systems, supporting defense against foreign DNA and facilitating nucleotide scavenging rather than direct antimicrobial resistance. This study provides new insights into the adaptive strategies of CPR bacteria and the multifunctional roles of MTs in metabolically streamlined microorganisms. ImportanceCandidate Phyla Radiation (CPR) microorganisms represent a major fraction of Earths microbial diversity, yet their biology remains poorly understood. We show that CPR genomes contain large numbers of methyltransferases and associated restriction enzymes, suggesting the existence of a specialized DNA-processing system. Because CPR organisms have highly reduced metabolic capabilities, this system may help them recover valuable nucleotides from external DNA while simultaneously protecting their genomes from foreign genetic material. We also found that CPR enzymes are highly distinct from those described in other bacteria and display remarkable functional diversity, highlighting the unique evolutionary trajectories of these microorganisms. By providing the first large-scale characterization of these enzymes in CPR, our study offers new insights into how these widespread microbes survive, acquire resources, and interact with their environment, expanding our understanding of microbial diversity and adaptation.

genomics↗

bioRxiv2020

Tricarboxylic acid cycle and proton gradient in Pandoravirus massiliensis: Is it still a virus?

Since the discovery of Acanthamoeba polyphaga Mimivirus, the first giant virus of amoeba, the historical hallmarks defining a virus have been challenged. Giant virion sizes can reach up to 2.3 {micro}m, making them visible by optical microscopy. They have large genomes of up to 2.5 Mb that encode proteins involved in the translation apparatus. Herein, we investigated possible energy production in Pandoravirus massiliensis, the largest of our giant virus collection. MitoTracker and TMRM mitochondrial membrane markers allowed for the detection of a membrane potential in virions that could be abolished by the use of the depolarizing agent CCCP. An attempt to identify enzymes involved in energy metabolism revealed that 8 predicted proteins of P. massiliensis exhibited low sequence identities with defined proteins involved in the universal tricarboxylic acid cycle (acetyl Co-A synthase; citrate synthase; aconitase; isocitrate dehydrogenase; -ketoglutarate decarboxylase; succinate dehydrogenase; fumarase). All 8 viral predicted ORFs were transcribed together during viral replication, mainly at the end of the replication cycle. Two of these proteins were detected in mature viral particles by proteomics. The product of the ORF132, a predicted protein of P. massiliensis, cloned and expressed in Escherichia coli, provided a functional isocitrate dehydrogenase, a key enzyme of the tricarboxylic acid cycle, which converts isocitrate to -ketoglutarate. We observed that membrane potential was enhanced by low concentrations of Acetyl-CoA, a regulator of the tricarboxylic acid cycle. Our findings show for the first time that energy production can occur in viruses, namely, pandoraviruses, and the involved enzymes are related to tricarboxylic acid cycle enzymes. The presence of a proton gradient in P. massiliensis coupled with the observation of genes of the tricarboxylic acid cycle make this virus a form a life for which it is legitimate to question what is a virus?.

microbiology↗

bioRxiv2019

A metallo-beta-lactamase with both beta-lactamase and ribonuclease activity is linked with traduction in giant viruses

Enzymatic proteins with a metallo-beta-lactamase (MBL) fold have been essentially studied in bacteria for their activity on beta-lactam antibiotics. However, the MBL fold is ancient and highly conserved, and these proteins are capable of cleaving a broad range of substrates. It has recently been shown that MBLs are present in a wide array of cellular organisms, including eukaryotes and archaea. We show here that Tupanvirus deep ocean, a giant virus, also encodes a protein with a MBL fold. Phylogeny showed its clustering with transfer ribonucleases (RNases) and the presence of orthologs in other giant viruses, mainly those harboring the largest sets of translation components. In addition, it suggests an ancient origin for these genes and a transfer between giant viruses and Acanthamoeba spp., a host of many giant viruses. Biologically, after its expression in Escherichia coli, the tupanvirus protein was found to hydrolyse nitrocefin, a chromogenic beta-lactam. We also observed an hydrolysis of penicillin G (10 g/mL) and detected the metabolite of penicillin G hydrolysis, benzylpenilloic acid. This was inhibited by sulbactam, a beta-lactamase inhibitor. In addition, we tested the degradation of single-stranded DNA, double-stranded DNA, and RNAs, and observed a strong activity on RNAs from seven bacteria with G+C varying from 42% to 67%, and from Acanthamoeba castellanii, the tupanvirus host. This was not inhibited by sulbactam or ceftriaxone. RNase activity was estimated to be 0.45{+/-}0.15 mU/mg using a fluorescence-based assay. Our results still broaden the range of hosts of MBL fold proteins and demonstrate that such protein can have dual beta-lactamase/nuclease activities. We suggest that they should be annotated according to this finding to avoid further confusion.

microbiology↗

bioRxiv2019

Paradoxical β-lactamase activity of archaeal encoding enzymes

{beta}-lactams targeting the bacterial cell wall are not active on archaea. Here, we figure out that annotation of genes as {beta}-lactamase in Archeae on the basis of homologous genes, initially annotated {beta}-lactamases, is a remnant of the identification of the original activities of this group of enzymes, which in fact, have multiple functions including nuclease, ribonuclease, {beta}-lactamase, or glyoxalase; which may specialized over time. We expressed a class B {beta}-lactamase enzyme from Methanosarcina barkeri that digest penicillin G. Moreover, while a weak glyoxalase activity was detected, a significant ribonuclease activity on bacterial and synthetic RNAs was demonstrated. The {beta}-lactamase activity was inhibited by a {beta}-lactamase inhibitor (sulbactam), but its RNAse activity was not. This gene appears to has been transferred to the Flavobacteriaceae group including Elizabethkingia genus in which the expressed gene shows a more specialized activity toward resistance to tienanmicin but no glyoxalase activity. The expressed class C-like {beta}-lactamase gene, also from Methanosarcina sp., shows also hydrolysis activity and was more closely related to DD-peptidase enzymes than known bacterial class C {beta}-lactamases. Our findings highlight the requalification needness of annotated enzymes as {beta}-lactamases and the specification overtime of multipotent enzymes in different ways in Archaea and bacteria.

microbiology↗