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Optimization and maintenance of soluble methane monooxygenase activity inMethylosinus trichosporium OB3b

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Abstract

Soluble methane monooxygenase (sMMO) maximization studies were carried out as part of a larger effort directed towards the development and optimization of an aqueous phase, multistage, membrane bioreactor system for treatment of polluted groundwater. A modified version of the naphthalene oxidation assay was utilized to determine the effects of methane:oxygen ratio, nutrient supply, and supplementary carbon sources on maximizing and maintaining sMMO activity inMethylosinus trichosporium OB3b.Methylosinus trichosporium OB3b attained peak sMMO activity (275–300 nmol of naphthol formed h−1 mg of protein−1 at 25°C) in early stationary growth phase when grown in nitrate mineral salts (NMS) medium. With the onset of methane limitation however, sMMO activity rapidly declined. It was possible to define a simplified nitrate mineral salts (NMS) medium, containing nitrate, phosphate and a source of iron and magnesium, which allowed reasonably high growth rates (μmax 0.08 h−1) and growth yields (0.4–0.5 g cells/g CH4) and near maximal activities of sMMO. In long term batch culture incubations sMMO activity reached a stable plateau at approximately 45–50% of the initial peak level and this was maintained over several weeks. The addition of d-biotin, pyridoxine, and vitamin B12 (cyanocobalamin) increased the activity level of sMMO in actively growing methanotrophs by 25–75%. The addition of these growth factors to the simplified NMS medium was found to increase the plateau sMMO level in long term batch cultures up to 70% of the original peak activity.

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Abbreviations

sMMO:

soluble methane monooxygenase

pMMO:

particulate methane monooxygenase

NMS:

nitrate mineral salts

TCE:

trichloroethene

NADH:

reduced nicotinamide adenine dinucleotide

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Authors and Affiliations

  1. Center for Environmental Biotechnology, The University of Tennessee, 37932, Knoxville, TN, USA

    John P. Bowman & Gary S. Sayler

  2. Graduate Program in Ecology, The University of Tennessee, 37932, Knoxville, TN, USA

    Gary S. Sayler

  3. Department of Microbiology, The University of Tennessee, 37932, Knoxville, TN, USA

    Gary S. Sayler

Authors
  1. John P. Bowman
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  2. Gary S. Sayler
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Bowman, J.P., Sayler, G.S. Optimization and maintenance of soluble methane monooxygenase activity inMethylosinus trichosporium OB3b. Biodegradation 5, 1–11 (1994). https://doi.org/10.1007/BF00695208

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  • DOI: https://doi.org/10.1007/BF00695208

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