Methane doubly-substituted isotopologue signatures (Δ13CH3D and Δ12CH2D2) of aerobic methane oxidation by Methylosinus trichosporium

ABSTRACT Two methanotrophic bacteria, Methylobacter albus BG8 and Methylosinus trichosporium OB3b, oxidized atmospheric methane during batch growth on methanol. Methane consumption was rapidly and substantially diminished (95% over 9 days) when washed cell suspensions were incubated without methanol in the presence of atmospheric methane (1.7 ppm). Methanotrophic activity was stimulated after methanol (10 mM) but not methane (1,000 ppm) addition. M. albus BG8 grown in continuous culture for 80 days with methanol retained the ability to oxidize atmospheric methane and oxidized methane in a chemostat air supply. Methane oxidation during growth on methanol was not affected by methane deprivation. Differences in the kinetics of methane uptake (apparent Km andV max) were observed between batch- and chemostat-grown cultures. The V max and apparent Km values (means ± standard errors) for methanol-limited chemostat cultures were 133 ± 46 nmol of methane 108 cells−1 h−1and 916 ± 235 ppm of methane (1.2 μM), respectively. These values were significantly lower than those determined with batch-grown cultures (V max of 648 ± 195 nmol of methane 108 cells−1 h−1 and apparent Km of 5,025 ± 1,234 ppm of methane [6.3 μM]). Methane consumption by soils was stimulated by the addition of methanol. These results suggest that methanol or other nonmethane substrates may promote atmospheric methane oxidation in situ.

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Dichloromethane and trichloroethylene inhibition of methane oxidation by the membrane-associated methane monooxygenase of Methylosinus trichosporium OB3b

Archives of Microbiology ◽

10.1007/s002030050714 ◽

1999 ◽

Vol 171 (5) ◽

pp. 301-308 ◽

Cited By ~ 13

Author(s):

Sonny Lontoh ◽

Alan A. DiSpirito ◽

J. D. Semrau

Keyword(s):

Methane Oxidation ◽

Methane Monooxygenase ◽

Methylosinus Trichosporium Ob3b ◽

Methylosinus Trichosporium

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Effect of Selected Monoterpenes on Methane Oxidation, Denitrification, and Aerobic Metabolism by Bacteria in Pure Culture

Applied and Environmental Microbiology ◽

10.1128/aem.64.2.520-525.1998 ◽

1998 ◽

Vol 64 (2) ◽

pp. 520-525 ◽

Cited By ~ 58

Author(s):

J. A. Amaral ◽

A. Ekins ◽

S. R. Richards ◽

R. Knowles

Keyword(s):

Methane Oxidation ◽

Optical Density ◽

Inhibitory Effect ◽

Aerobic Metabolism ◽

Environmental Isolates ◽

Methylosinus Trichosporium ◽

Factors Affecting ◽

Pure Cultures ◽

Limonene Oxide ◽

Significant Factors

ABSTRACT Selected monoterpenes inhibited methane oxidation by methanotrophs (Methylosinus trichosporium OB3b, Methylobacter luteus), denitrification by environmental isolates, and aerobic metabolism by several heterotrophic pure cultures. Inhibition occurred to various extents and was transient. Complete inhibition of methane oxidation by Methylosinus trichosporium OB3b with 1.1 mM (−)-α-pinene lasted for more than 2 days with a culture of optical density of 0.05 before activity resumed. Inhibition was greater under conditions under which particulate methane monooxygenase was expressed. No apparent consumption or conversion of monoterpenes by methanotrophs was detected by gas chromatography, and the reason that transient inhibition occurs is not clear. Aerobic metabolism by several heterotrophs was much less sensitive than methanotrophy was;Escherichia coli (optical density, 0.01), for example, was not affected by up to 7.3 mM (−)-α-pinene. The degree of inhibition was monoterpene and species dependent. Denitrification by isolates from a polluted sediment was not inhibited by 3.7 mM (−)-α-pinene, γ-terpinene, or β-myrcene, whereas 50 to 100% inhibition was observed for isolates from a temperate swamp soil. The inhibitory effect of monoterpenes on methane oxidation was greatest with unsaturated, cyclic hydrocarbon forms [e.g., (−)-α-pinene, (S)-(−)-limonene, (R)-(+)-limonene, and γ-terpinene]. Lower levels of inhibition occurred with oxide and alcohol derivatives [(R)-(+)-limonene oxide, α-pinene oxide, linalool, α-terpineol] and a noncyclic hydrocarbon (β-myrcene). Isomers of pinene inhibited activity to different extents. Given their natural sources, monoterpenes may be significant factors affecting bacterial activities in nature.

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Genome Sequence of the Obligate Methanotroph Methylosinus trichosporium Strain OB3b

Journal of Bacteriology ◽

10.1128/jb.01144-10 ◽

2010 ◽

Vol 192 (24) ◽

pp. 6497-6498 ◽

Cited By ~ 59

Author(s):

Lisa Y. Stein ◽

Sukhwan Yoon ◽

Jeremy D. Semrau ◽

Alan A. DiSpirito ◽

Andrew Crombie ◽

...

Keyword(s):

Methane Oxidation ◽

Genome Sequence ◽

Catalytic Properties ◽

Methane Monooxygenase ◽

Structure And Function ◽

Methylosinus Trichosporium ◽

Soluble Methane Monooxygenase ◽

Obligate Aerobic ◽

Copper Chelator ◽

And Function

ABSTRACT Methylosinus trichosporium OB3b (for “oddball” strain 3b) is an obligate aerobic methane-oxidizing alphaproteobacterium that was originally isolated in 1970 by Roger Whittenbury and colleagues. This strain has since been used extensively to elucidate the structure and function of several key enzymes of methane oxidation, including both particulate and soluble methane monooxygenase (sMMO) and the extracellular copper chelator methanobactin. In particular, the catalytic properties of soluble methane monooxygenase from M. trichosporium OB3b have been well characterized in context with biodegradation of recalcitrant hydrocarbons, such as trichloroethylene. The sequence of the M. trichosporium OB3b genome is the first reported from a member of the Methylocystaceae family in the order Rhizobiales.

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Methane Monooxygenase Gene Transcripts as Quantitative Biomarkers of Methanotrophic Activity in Methylosinus trichosporium OB3b

Applied and Environmental Microbiology ◽

10.1128/aem.01048-20 ◽

2020 ◽

Vol 86 (23) ◽

Author(s):

Egidio F. Tentori ◽

Ruth E. Richardson

Keyword(s):

Methane Oxidation ◽

Methane Monooxygenase ◽

Methane Emissions ◽

Mrna Transcript ◽

Methylosinus Trichosporium ◽

Transcript Levels ◽

Content Type ◽

Oxidation Rates ◽

Pure Cultures

ABSTRACT Methanotrophic microorganisms are characterized by their ability to oxidize methane. Globally they have a significant impact on methane emissions by attenuating net methane fluxes to the atmosphere in natural and engineered systems, though the populations are dynamic in their activity level in soils and waters. Methanotrophs oxidize methane using methane monooxygenase (MMO) enzymes, and selected subunit genes of the most common MMOs, specifically pmoA and mmoX, are used as biomarkers for the presence and abundance of populations of bacterial methanotrophs. The relative expression of these biomarker genes is dependent on copper-to-biomass ratios. Empirically derived quantitative relationships between methane oxidation biomarker transcript amounts and methanotrophic activity could facilitate determination of methane oxidation rates. In this study, pure cultures of a model type II methanotroph, Methylosinus trichosporium OB3b, were grown in hollow-fiber membrane bioreactors (HFMBR) under different steady-state methane oxidation conditions. Methanotroph biomass (DNA based) and methane oxidation biomarker mRNA transcript amounts were determined using quantitative PCR (qPCR) and reverse transcription-PCR (RT-qPCR), respectively. Under both copper-present and copper-limited conditions, per-cell pmoA mRNA transcript levels positively correlated with measured per-cell methane oxidation rates across 3 orders of magnitude. These correlations, if maintained across different methanotrophs, could prove valuable for inferring in situ oxidation rates of methanotrophs and understanding the dynamics of their impact on net methane emissions. IMPORTANCE Methanotrophs are naturally occurring microorganisms capable of oxidizing methane and have an impact on global net methane emissions. The genes pmoA and mmoX are used as biomarkers for bacterial methanotrophs. Quantitative relationships between transcript amounts of these genes and methane oxidation rates could facilitate estimation of methanotrophic activity. In this study, a strong correlation was observed between per-cell pmoA transcript levels and per-cell methane oxidation rates for pure cultures of the aerobic methanotroph M. trichosporium OB3b grown in bioreactors. If similar relationships exist across different methanotrophs, they could prove valuable for inferring in situ oxidation rates of methanotrophs and better understanding their impact on net methane emissions.

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Biosynthesis of methanol from methane by Methylosinus trichosporium OB3b

Chemical Papers ◽

10.2478/s11696-008-0100-5 ◽

2009 ◽

Vol 63 (2) ◽

Cited By ~ 12

Author(s):

Agata Markowska ◽

Beata Michalkiewicz

Keyword(s):

Methane Oxidation ◽

Bacterial Biomass ◽

Step Method ◽

Methylosinus Trichosporium Ob3b ◽

Methylosinus Trichosporium ◽

Methanol Production ◽

Whole Cells ◽

Conversion Of Methane ◽

One Step ◽

The One

AbstractMethanol has recently attracted significant interest in the energetic field. Current technology for the conversion of methane to methanol is based on energy intensive endothermic steam reforming followed by catalytic conversion into methanol. The one-step method performed at very low temperatures (35°C) is methane oxidation to methanol via bacteria. The aim of this work was to examine the role of copper in the one-step methane oxidation to methanol by utilizing whole cells of Methylosinus trichosporium OB3b bacteria. From the results obtained it was found that copper concentration in the medium influences the rate of bacterial biomass growth or methanol production during the process of methane oxidation to methanol. The presented results indicate that the process of methane oxidation to methanol by Methylosinus trichosporium OB3b bacteria is most efficient when the mineral medium contains 1.0 × 10−6 mol dm−3 of copper. Under these conditions, a satisfactory growth of biomass was also achieved.

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Ammonium and Nitrite Inhibition of Methane Oxidation by Methylobacter albus BG8 and Methylosinus trichosporium OB3b at Low Methane Concentrations †

Applied and Environmental Microbiology ◽

10.1128/aem.60.10.3508-3513.1994 ◽

1994 ◽

Vol 60 (10) ◽

pp. 3508-3513 ◽

Cited By ~ 103

Author(s):

Gary M. King ◽

Sylvia Schnell

Keyword(s):

Methane Oxidation ◽

Methylosinus Trichosporium Ob3b ◽

Methylosinus Trichosporium ◽

Nitrite Inhibition ◽

Methane Concentrations

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Genome-Scale Metabolic Model Reconstruction and in Silico Investigations of Methane Metabolism in Methylosinus trichosporium OB3b

Microorganisms ◽

10.3390/microorganisms8030437 ◽

2020 ◽

Vol 8 (3) ◽

pp. 437 ◽

Cited By ~ 4

Author(s):

Sanzhar Naizabekov ◽

Eun Yeol Lee

Keyword(s):

Methane Oxidation ◽

In Silico ◽

Model Organism ◽

Metabolic Model ◽

Central Carbon Metabolism ◽

Methylosinus Trichosporium Ob3b ◽

Methylosinus Trichosporium ◽

Specific Growth Rates ◽

Methane Metabolism ◽

Genome Scale

Methylosinus trichosporium OB3b is an obligate aerobic methane-utilizing alpha-proteobacterium. Since its isolation, M. trichosporium OB3b has been established as a model organism to study methane metabolism in type II methanotrophs. M. trichosporium OB3b utilizes soluble and particulate methane monooxygenase (sMMO and pMMO respectively) for methane oxidation. While the source of electrons is known for sMMO, there is less consensus regarding electron donor to pMMO. To investigate this and other questions regarding methane metabolism, the genome-scale metabolic model for M. trichosporium OB3b (model ID: iMsOB3b) was reconstructed. The model accurately predicted oxygen: methane molar uptake ratios and specific growth rates on nitrate-supplemented medium with methane as carbon and energy source. The redox-arm mechanism which links methane oxidation with complex I of electron transport chain has been found to be the most optimal mode of electron transfer. The model was also qualitatively validated on ammonium-supplemented medium indicating its potential to accurately predict methane metabolism in different environmental conditions. Finally, in silico investigations regarding flux distribution in central carbon metabolism of M. trichosporium OB3b were performed. Overall, iMsOB3b can be used as an organism-specific knowledgebase and a platform for hypothesis-driven theoretical investigations of methane metabolism.

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