Effect of Growth Conditions on the Expression of Soluble Methane Monooxygenase

Ana Begonja and Dubravka Hršak*

Center for Marine and Environmental Research, Ruđer Bošković Institute, POB 180, HR-10002 Zagreb, Croatia

Article history:
Received May 3, 2000
Accepted October 26, 2000

Key words:
soluble methane monooxygenase, type II methanotroph, methanotrophic-heterotrophic community

Summary:
Methanotrophic-heterotrophic groundwater community (culture MM1) and the obligate type II methanotroph (strain Met1) isolated from the culture MM1 were found to express soluble methane monooxygenase (sMMO) enzyme system when grown in simple batch culture (shake flasks) under the determined conditions. To optimize conditions for the enzyme expression, effects of the essential constituents of nitrate mineral salts (NMS) medium on the growth and sMMO activity was studied. The obtained results confirmed that copper deficiency was essential for sMMO expression in both cases, i.e. when methanotrophic strain Met1 was grown in the community and as a single culture. Furthermore, under the experimental conditions the specific whole-cell sMMO activity of the strain Met1, measured by the naphthalene oxidation assay, showed no significant effect on the increased KNO₃ concentration from 10 to 20 mM. In contrast, when KNO₃ was replaced by NH₄Cl, both the growth of the strain Met1 and the sMMO activity were highly limited. The significant positive effect on the growth and sMMO activity was observed by increasing the initial FeSO₄ concentration in NMS medium from 12 to 52
µM. This suggested that Fe²⁺ is an essential medium constituent, which can contribute the most to the optimization of shake flask system for sMMO expression.

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