biostudies-literatureHurt AGreat Lakes Bioenergy Research Center, U.S. Department of Energy, Office of Science, Office of Biological and Environmental ResearchUnited States Department of EnergyDepartment of Biochemistry and Molecular Biology and startup funding and support from AgBioResearch500https://www.ebi.ac.uk/biostudies/studies/S-EPMC10974752biostudies-literatureUnknown12(3)Terpenes are diverse specialized metabolites naturally found within plants and have important roles in inter-species communication, adaptation and interaction with the environment. Their industrial applications span a broad range, including fragrances, flavors, cosmetics, natural colorants to agrochemicals and therapeutics, yet formal chemical synthesis is economically challenging due to structural complexities. Engineering terpene biosynthesis could represent an alternative in microbial biotechnological workhorses, such as <i>Saccharomyces cerevisiae</i> or <i>Escherichi coli</i>, utilizing sugars or complex media as feedstocks. Host species that metabolize renewable and affordable carbon sources may offer unique sustainable biotechnological alternatives. Methylotrophs are bacteria with the capacity to utilize one-carbon feedstocks, such as methanol or formate. They colonize the phyllosphere (above-ground area) of plants, and many accumulate abundant carotenoid pigments. Methylotrophs have the capacity to take up and use a subset of the rare earth elements known as lanthanides. These metals can enhance one-carbon (methylotrophic) metabolism. Here, we investigated whether manipulating the metabolism enables and enhances terpene production. A carotenoid-deficient mutant potentially liberates carbon, which may contribute to bioproduct accumulation. To test this hypothesis, terpene-producing bacterial strains regulated by two distinct promoters were generated. Wildtype <i>Methylobacterium extorquens</i>, ∆<i>Meta1_3665</i>, a methylotrophic mutant lacking the carotenoid pathway, and an <i>E. coli</i> strain were transformed with an exogenous terpene pathway and grown both in the presence and absence of lanthanides. The extraction, and the comparison of analytical profiles, provided evidence that engineered cultured <i>M. extorquens</i> under control of a native, inducible methylotrophic promoter can yield the sesquiterpene patchoulol when supplemented with lanthanide. In contrast, using a moderate-strength constitutive promoter failed to give production. We demonstrated colonization of the phyllosphere with the engineered strains, supporting the future engineering of selected species of the plant microbiome and with promising implications for the synthetic biology of small molecules.MicroorganismsEngineering Terpene Production Pathways in <i>Methylobacterium extorquens</i> AM1.PMC10974752MICL02454DE-SC0018409Bibik JDHurt AMartinez-Gomez NCHamberger BfalseEngineering Terpene Production Pathways in <i>Methylobacterium extorquens</i> AM1.Terpenes are diverse specialized metabolites naturally found within plants and have important roles in inter-species communication, adaptation and interaction with the environment. Their industrial applications span a broad range, including fragrances, flavors, cosmetics, natural colorants to agrochemicals and therapeutics, yet formal chemical synthesis is economically challenging due to structural complexities. Engineering terpene biosynthesis could represent an alternative in microbial biotechnological workhorses, such as <i>Saccharomyces cerevisiae</i> or <i>Escherichi coli</i>, utilizing sugars or complex media as feedstocks. Host species that metabolize renewable and affordable carbon sources may offer unique sustainable biotechnological alternatives. Methylotrophs are bacteria with the capacity to utilize one-carbon feedstocks, such as methanol or formate. They colonize the phyllosphere (above-ground area) of plants, and many accumulate abundant carotenoid pigments. Methylotrophs have the capacity to take up and use a subset of the rare earth elements known as lanthanides. These metals can enhance one-carbon (methylotrophic) metabolism. Here, we investigated whether manipulating the metabolism enables and enhances terpene production. A carotenoid-deficient mutant potentially liberates carbon, which may contribute to bioproduct accumulation. To test this hypothesis, terpene-producing bacterial strains regulated by two distinct promoters were generated. Wildtype <i>Methylobacterium extorquens</i>, ∆<i>Meta1_3665</i>, a methylotrophic mutant lacking the carotenoid pathway, and an <i>E. coli</i> strain were transformed with an exogenous terpene pathway and grown both in the presence and absence of lanthanides. The extraction, and the comparison of analytical profiles, provided evidence that engineered cultured <i>M. extorquens</i> under control of a native, inducible methylotrophic promoter can yield the sesquiterpene patchoulol when supplemented with lanthanide. In contrast, using a moderate-strength constitutive promoter failed to give production. We demonstrated colonization of the phyllosphere with the engineered strains, supporting the future engineering of selected species of the plant microbiome and with promising implications for the synthetic biology of small molecules.2024-01-01T00:00:00Z2024 Feb2026-04-08T15:26:11.358Z2025-04-05T23:15:56.332ZS-EPMC109747523854355110.3390/microorganisms12030500