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Variation of salinity and nitrogen concentration affects the pentacyclic triterpenoid inventory of the haloalkaliphilic aerobic methanotrophic bacterium Methylotuvimicrobium alcaliphilum.

ABSTRACT: The occurrence and activity of aerobic methanotrophs are influenced by environmental conditions, including pH, temperature, salinity, methane and oxygen concentrations, and nutrient availability. Aerobic methanotrophs synthesize a variety of lipids important for cell functions. However, culture-based experiments studying the influence of environmental parameters on lipid production by aerobic methanotrophs are scarce. Such information is crucial to interpret lipid patterns of methanotrophic bacteria in the environment. In this study, the alkaliphilic strain Methylotuvimicrobium alcaliphilum was cultivated under different salinities and different nitrate concentrations to assess the effect of changing conditions on the inventory of pentacyclic triterpenoids. The results indicate that hopanoid abundance is enhanced at lower salinity and higher nitrate concentration. The production of most pentacyclic triterpenoids was favored at low salinity, especially for aminotriol. Interestingly, 3-methyl-aminotetrol and tetrahymanol were favored at higher salinity. Bacteriohopanepolyols (BHPs), particularly aminotriol and 3-methyl-aminotriol, increased considerably at higher nitrate concentrations. Four novel N-containing BHPs-aminodiol, 3-methyl-aminodiol, and isomers of aminotriol and 3-methyl-aminotriol-were identified. This study highlights the significance of environmental factors for bacterial lipid production and documents the need for cultivation studies under variable conditions to utilize the full potential of the biomarker concept.

SUBMITTER: Cordova-Gonzalez A

PROVIDER: S-EPMC8102298 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:BackgroundMethane is the primary component of natural gas and biogas. The huge abundance of methane makes it a promising alternative carbon source for industrial biotechnology. Herein, we report diamine compound, putrescine, production from methane by an industrially promising methanotroph Methylomicrobium alcaliphilum 20Z.ResultsWe conducted adaptive evolution to improve putrescine tolerance of M. alcaliphilum 20Z because putrescine highly inhibits the cell growth. The evolved strain 20ZE was able to grow in the presence of 400 mM of putrescine dihydrochloride. The expression of linear pathway ornithine decarboxylase genes from Escherichia coli and Methylosinus trichosporium OB3b allowed the engineered strain to produce putrescine. A higher putrescine titer of 12.44 mg/L was obtained in the strain 20ZE-pACO with ornithine decarboxylase from M. trichosporium OB3b. For elimination of the putrescine utilization pathway, spermidine synthase (MEALZ_3408) was knocked out, resulting in no spermidine formation in the strain 20ZES1-pACO with a putrescine titer of 18.43 mg/L. Next, a genome-scale metabolic model was applied to identify gene knockout strategies. Acetate kinase (MEALZ_2853) and subsequently lactate dehydrogenase (MEALZ_0534) were selected as knockout targets, and the deletion of these genes resulted in an improvement of the putrescine titer to 26.69 mg/L. Furthermore, the putrescine titer was improved to 39.04 mg/L by overexpression of key genes in the ornithine biosynthesis pathway under control of the pTac promoter. Finally, suitable nitrogen sources for growth of M. alcaliphilum 20Z and putrescine production were optimized with the supplement of 2 mM ammonium chloride to nitrate mineral salt medium, and this led to the production of 98.08 mg/L putrescine, almost eightfold higher than that from the initial strain. Transcriptome analysis of the engineered strains showed upregulation of most genes involved in methane assimilation, citric acid cycle, and ammonia assimilation in ammonia nitrate mineral salt medium, compared to nitrate mineral salt medium.ConclusionsThe engineered M. alcaliphilum 20ZE4-pACO strain was able to produce putrescine up to 98.08 mg/L, almost eightfold higher than the initial strain. This study represents the bioconversion of methane to putrescine-a high value-added diamine compound.

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Variation of salinity and nitrogen concentration affects the pentacyclic triterpenoid inventory of the haloalkaliphilic aerobic methanotrophic bacterium Methylotuvimicrobium alcaliphilum.

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