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1. Aromatic acid metabolism in Methylobacterium extorquens reveals interplay between methylotrophic and heterotrophic pathways

   https://doi.org/10.1128/aem.00761-25  

   Govindaraju, Alekhya M; Martinez-Gomez, Norma Cecilia (September 2025, Applied and Environmental Microbiology)

   Bose, Arpita (Ed.)

   ABSTRACT Efforts toward microbial conversion of lignin to value-added products face many challenges because lignin’s methoxylated aromatic monomers release toxic C₁byproducts such as formaldehyde. The ability to grow on methoxylated aromatic acids (e.g., vanillic acid) has been identified in certain clades of methylotrophs, bacteria characterized by their unique ability to tolerate and metabolize high concentrations of formaldehyde. Here, we use a phyllosphere methylotroph isolate,Methylobacterium extorquensSLI 505, as a model to identify the fate of formaldehyde during methylotrophic growth on vanillic acid.M. extorquensSLI 505 displays concentration-dependent growth phenotypes on vanillic acid without concomitant formaldehyde accumulation. We conclude thatM. extorquensSLI 505 overcomes metabolic bottlenecks from simultaneous assimilation of multicarbon and C₁intermediates by allocating formaldehyde toward dissimilation and assimilating the ring carbons of vanillic acid heterotrophically. We correlate this strategy with maximization of bioenergetic yields and demonstrate that formaldehyde dissimilation for energy generation rather than formaldehyde detoxification is advantageous for growth on aromatic acids.M. extorquensSLI 505 also exhibits catabolite repression during growth on methanol and low concentrations of vanillic acid, but no diauxic patterns during growth on methanol and high concentrations of vanillic acid. Results from this study outline metabolic strategies employed byM. extorquensSLI 505 for growth on a complex single substrate that generates both C₁and multicarbon intermediates and emphasizes the robustness ofM. extorquensfor biotechnological applications for lignin valorization.IMPORTANCELignin, one of the most abundant and renewable carbon sources on Earth, is a promising alternative to non-renewable fossil fuels used to produce petrochemicals. Degradation of lignin releases toxic C₁byproducts such as formaldehyde, and thus most microorganisms are not suitable for biorefining lignin. By contrast,Methylobacterium extorquensSLI 505 is capable of growth on high concentrations of aromatic acids without concomitant formaldehyde accumulation. In addition, we show that the growth ofM. extorquensSLI 505 on aromatic acids is coupled to the production of the bioplastic, polyhydroxybutyrate. Aromatic acids serve as a model by which to understand howM. extorquensSLI 505 balances methylotrophic and heterotrophic pathways during growth to provide strategies for growth optimization when using complex substrates in both ecological and industrial fermentation applications. 

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2. 5–5 Lignin Linkage Cleavage over Ru: A Density Functional Theory Study

   https://doi.org/10.1021/acssuschemeng.1c04838  

   Li, Qiang; Vlachos, Dionisios G (December 2021, ACS Sustainable Chemistry & Engineering)

   Lignin is the most abundant natural, aromatic-containing biopolymer. Among all the C–C and C–O bonds being cleaved in catalytic fractionation, the 5–5 linkage is the strongest, and its scission requires harsh conditions. Theoretical investigations of the mechanism and kinetics could provide insights into developing better catalysts but are essentially lacking. We perform extensive density functional theory calculations on 2-methoxy-1,1′-biphenyl, a model compound, with various substitutions at all ring locations on Ru(0001). We analyze the competition between the 5–5 bond cleavage and the defunctionalization of the side functional groups at multiple degrees of depolymerization. The role of ring functional groups in the adsorption of lignin oligomers and the 5–5 bond scission and, conversely, the effect of the aromatic group on the −OCH3 decomposition are also discussed. We show that increasing the number of methoxy groups decreases the C–C barrier, and thus, we expect the following depolymerization ranking: grass > softwood > hardwood. While Ru exposes modest 5–5 bond scission reaction barriers from some intermediates, rapid side group chemistry prevents the formation of these intermediates; instead, scission happens most probably from defunctionalized compounds whose C–C scission barriers are high. Our results also expose the existence of multiple Brønsted–Evans–Polanyi relations in the catalytic transformation of biphenyl-based molecules that open up the possibility of modeling depolymerization of large lignin chains. 

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3. Enabling microbial syringol conversion through structure-guided protein engineering

   https://doi.org/10.1073/pnas.1820001116  

   Machovina, Melodie M.; Mallinson, Sam J.; Knott, Brandon C.; Meyers, Alexander W.; Garcia-Borràs, Marc; Bu, Lintao; Gado, Japheth E.; Oliver, April; Schmidt, Graham P.; Hinchen, Daniel J.; et al (January 2019, Proceedings of the National Academy of Sciences)

   Microbial conversion of aromatic compounds is an emerging and promising strategy for valorization of the plant biopolymer lignin. A critical and often rate-limiting reaction in aromatic catabolism is O -aryl-demethylation of the abundant aromatic methoxy groups in lignin to form diols, which enables subsequent oxidative aromatic ring-opening. Recently, a cytochrome P450 system, GcoAB, was discovered to demethylate guaiacol (2-methoxyphenol), which can be produced from coniferyl alcohol-derived lignin, to form catechol. However, native GcoAB has minimal ability to demethylate syringol (2,6-dimethoxyphenol), the analogous compound that can be produced from sinapyl alcohol-derived lignin. Despite the abundance of sinapyl alcohol-based lignin in plants, no pathway for syringol catabolism has been reported to date. Here we used structure-guided protein engineering to enable microbial syringol utilization with GcoAB. Specifically, a phenylalanine residue (GcoA-F169) interferes with the binding of syringol in the active site, and on mutation to smaller amino acids, efficient syringol O -demethylation is achieved. Crystallography indicates that syringol adopts a productive binding pose in the variant, which molecular dynamics simulations trace to the elimination of steric clash between the highly flexible side chain of GcoA-F169 and the additional methoxy group of syringol. Finally, we demonstrate in vivo syringol turnover in Pseudomonas putida KT2440 with the GcoA-F169A variant. Taken together, our findings highlight the significant potential and plasticity of cytochrome P450 aromatic O -demethylases in the biological conversion of lignin-derived aromatic compounds. 

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4. EfgA is a conserved formaldehyde sensor that leads to bacterial growth arrest in response to elevated formaldehyde

   https://doi.org/doi: 10.131/journal.pbio.3001208  

   Bazurto, Jannell V.; Nayak, Dipiti; Ticak, Tomislav; Davlieva, Milya; Lee, Jessica A.; Hellenbrand, Chandler N.; Lambert, Leah B.; Benski, Oliva J.; Quates, Caleb J.; Johnson, Jill L.; et al (May 2021, PLoS biology)

   null (Ed.)

   Normal cellular processes give rise to toxic metabolites that cells must mitigate. Formaldehyde is a universal stressor and potent metabolic toxin that is generated in organisms from bacteria to humans. Methylotrophic bacteria such as Methylorubrum extorquens face an acute challenge due to their production of formaldehyde as an obligate central intermediate of single-carbon metabolism. Mechanisms to sense and respond to formaldehyde were speculated to exist in methylotrophs for decades but had never been discovered. Here, we identify a member of the DUF336 domain family, named efgA for enhanced formaldehyde growth, that plays an important role in endogenous formaldehyde stress response in M. extorquens PA1 and is found almost exclusively in methylotrophic taxa. Our experimental analyses reveal that EfgA is a formaldehyde sensor that rapidly arrests growth in response to elevated levels of formaldehyde. Heterologous expression of EfgA in Escherichia coli increases formaldehyde resistance, indicating that its interaction partners are widespread and conserved. EfgA represents the first example of a formaldehyde stress response system that does not involve enzymatic detoxification. Thus, EfgA comprises a unique stress response mechanism in bacteria, whereby a single protein directly senses elevated levels of a toxic intracellular metabolite and safeguards cells from potential damage. 

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5. Valorization of Lignin as a Sustainable Component of Structural Materials and Composites: Advances from 2011 to 2019

   https://doi.org/10.3390/su12020734  

   Karunarathna, Menisha S.; Smith, Rhett C. (January 2020, Sustainability)

   null (Ed.)

   Lignin is the most abundant aromatic biopolymer and is the sustainable feedstock most likely to supplant petroleum-derived aromatics and downstream products. Rich in functional groups, lignin is largely peerless in its potential for chemical modification towards attaining target properties. Lignin’s crosslinked network structure can be exploited in composites to endow them with remarkable strength, as exemplified in timber and other structural elements of plants. Yet lignin may also be depolymerized, modified, or blended with other polymers. This review focuses on substituting petrochemicals with lignin derivatives, with a particular focus on applications more significant in terms of potential commercialization volume, including polyurethane, phenol-formaldehyde resins, lignin-based carbon fibers, and emergent melt-processable waste-derived materials. This review will illuminate advances from the last eight years in the prospective utilization of such lignin-derived products in a range of application such as adhesives, plastics, automotive components, construction materials, and composites. Particular technical issues associated with lignin processing and emerging alternatives for future developments are discussed. 

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