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

Whole-cell assays were used to measure the effect of dichloromethane and tr ichloroethylene on methane oxidation by Methylosinus trichosporium OB3b syn thesizing the membrane-associated or particulate methane monooxygenase (pMM O). For M. trichosporium OB3b grown with 20 CIM copper, no inhibition of me thane oxidation was observed in the presence of either dichloromethane or t richloroethylene. If 20 mM formate was added to the reaction vials, however , methane oxidation rates increased and inhibition of methane oxidation was observed in the presence of dichloromethane and trichloroethylene. In the presence of formate, dichloromethane acted as a competitive inhibitor, whil e trichloroethylene acted as a noncompetitive inhibitor. The finding of non competitive inhibition by trichloroethylene was further examined by measuri ng the inhibition constants K-iE and K-iES. These constants suggest that tr ichloroethylene competes with methane at some sites, although it can bind t o others if methane is already bound. Whole-cell oxygen uptake experiments for active and acetylene-treated cells also showed that provision of format s could stimulate both methane and trichloroethylene oxidation and that tri chloroethylene did not affect formate dehydrogenase activity. The finding t hat different chlorinated hydrocarbons caused different inhibition patterns can be explained by either multiple substrate binding sites existing in pM MO or multiple forms of pMMO with different activities. The whole-cell anal ysis performed here cannot distinguish between these models, and further wo rk should be done on obtaining active preparations of the purified pMMO.