Bazurto, J. V. | Search bioRxiv

bioRxiv2026

Decreasing peptide deformylase activity is a beneficial strategy for increasing formaldehyde resistance in Methylobacterium extorquens

Formaldehyde is a highly toxic metabolite that can cause extensive damage to DNA and proteins, and strategies to mitigate formaldehyde toxicity are poorly understood. Methylotrophic bacteria, such as Methylobacterium extorquens, thrive on one-carbon compounds as sole sources of carbon and energy. These organisms are excellent models for discovering formaldehyde stress response systems because formaldehyde is an obligate intermediate in their central carbon metabolism. Here, we characterize an evolved def allele (defevo) that increases formaldehyde resistance in M. extorquens. The def gene encodes peptide deformylase (PDF, EC:3.5.1.88), an enzyme that contributes to protein processing by removing the formyl group from N-formylmethionine (fMet) on nascent peptides. The defevoallele has a single missense mutation that decreases PDF activity both in vitro and in vivo. Transcriptomic analysis of the defevo strain indicates there are pleiotropic effects of this mutation and a differential response to formaldehyde stress. We investigate possible mechanisms for the defevo mutants increased resistance to formaldehyde, including mitigation of formaldehyde-induced protein stress and altered membrane physiology. We find that the defevo allele selectively alleviates exogenous, but not endogenous, formaldehyde stress and identify a tradeoff in heat shock resistance. This study reports the first observation of lowered PDF activity benefiting a cellular physiological phenotype. Our work indicates that altered protein metabolism can mitigate the toxic effects of formaldehyde and furthers our understanding of the strategies that can protect cells from formaldehyde-induced damage. ImportanceFormaldehyde is a toxic chemical that can damage essential molecules inside of cells, yet all organisms inevitably produce it during normal metabolism. Despite its ubiquity, our understanding of strategies for how cells navigate formaldehyde toxicity is incomplete. This study focuses on Methylobacterium extorquens, which naturally generates high levels of formaldehyde as part of its growth on simple carbon compounds. We show herein that a single genetic change, which slows down how newly made proteins are processed during translation, can unexpectedly improve the bacteriums ability to resist formaldehyde stress. Further, we show that this single change has numerous effects on the cell, many of which may contribute to formaldehyde resistance.

physiology↗

bioRxiv2020

Formaldehyde-responsive proteins, TtmR and EfgA, reveal a tradeoff between formaldehyde resistance and efficient transition to methylotrophy in Methylorubrum extorquens

For bacteria to thrive they must be well-adapted to their environmental niche, which may involve specialized metabolism, timely adaptation to shifting environments, and/or the ability to mitigate numerous stressors. These attributes are highly dependent on cellular machinery that can sense both the external and intracellular environment. Methylorubrum extorquens is an extensively studied facultative methylotroph, an organism that can use single-carbon compounds as their sole source of carbon and energy. In methylotrophic metabolism, carbon flows through formaldehyde as a central metabolite; thus, formaldehyde is both an obligate metabolite and a metabolic stressor. Via the one-carbon dissimilation pathway, free formaldehyde is rapidly incorporated by formaldehyde activating enzyme (Fae), which is constitutively expressed at high levels. In the presence of elevated formaldehyde levels, a recently identified formaldehyde-sensing protein, EfgA, induces growth arrest. Herein, we describe TtmR, a formaldehyde-responsive transcription factor that, like EfgA, modulates formaldehyde resistance. TtmR is a member of the MarR family of transcription factors and impacts the expression of 75 genes distributed throughout the genome, many of which are transcription factors and/or involved in stress response, including efgA. Notably, when M. extorquens is adapting its metabolic network during the transition to methylotrophy, efgA and ttmR mutants experience an imbalance in formaldehyde production and a notable growth delay. Although methylotrophy necessitates that M. extorquens maintain a relatively high level of formaldehyde tolerance, this work reveals a tradeoff between formaldehyde resistance and the efficient transition to methylotrophic growth and suggests that TtmR and EfgA play a pivotal role in maintaining this balance. ImportanceAll organisms produce formaldehyde as a byproduct of enzymatic reactions and as a degradation product of metabolites. The ubiquity of formaldehyde in cellular biology suggests all organisms have evolved mechanisms of mitigating formaldehyde toxicity. However, formaldehyde-sensing is poorly described and prevention of formaldehyde-induced damage is primarily understood in the context of detoxification. Here we use an organism that is regularly exposed to elevated intracellular formaldehyde concentrations through high-flux one-carbon utilization pathways to gain insight into the role of formaldehyde-responsive proteins that modulate formaldehyde resistance. Using a combination of genetic and transcriptomic analyses, we identify dozens of genes putatively involved in formaldehyde resistance, determined the relationship between two different formaldehyde response systems and identified an inherent tradeoff between formaldehyde resistance and optimal transition to methylotrophic metabolism.

microbiology↗