Redox properties of the hydroxylase component of methane monooxygenase from Methylococcus capsulatus (Bath). Effects of protein B, reductase, and substrate.
The reduction potentials of the hydroxylase component of the soluble methane monooxygenase from Methylococcus capsulatus (Bath) have been investigated through potentiometric titrations. The potentials were determined by EPR spectroscopic quantitation of the mixed valent hydroxylase as a function of added sodium dithionite in the presence of appropriate mediators. The reduction of the oxidized Fe(III).Fe(III) form to the mixed valent Fe(II).Fe(III) form occurs at 48 mV versus NHE while the potential for the formation of the fully reduced Fe(II).Fe(II) species from the mixed valent form was determined to be -135 mV. Addition of the substrate propylene to the hydroxylase did not have a major effect on the reduction potentials. Introduction of the protein B and the reductase components, however, completely inhibited reduction of the hydroxylase at potentials as far negative as -200 mV. Addition of propylene to all three methane monooxygenase components greatly facilitated hydroxylase reduction. Under these conditions, the fully reduced form of the protein was obtained at potentials of greater than 150 mV. This high redox potential indicates that the oxidized form of the protein is highly reactive, as required for methane oxidation. The present results reveal aspects of how both protein B and substrate can regulate electron transfer into and out of the hydroxylase component of methane monooxygenase.
Liu
K E
KE
Department of Chemistry, Massachusetts Institute of Technology, Cambridge 02139.
Lippard
S J
SJ
D004227:Dithionite / Q000494:pharmacology; D004578:Electron Spin Resonance Spectroscopy; D019605:Euryarchaeota / Q000201:enzymology*; D007501:Iron / Q000378:metabolism; D007700:Kinetics; D006899:Mixed Function Oxygenases / Q000378:metabolism*; D009097:Multienzyme Complexes / Q000378:metabolism*; D010084:Oxidation-Reduction; D010105:Oxygenases / Q000378:metabolism*
D009097:Multienzyme Complexes; D004227:Dithionite; D007501:Iron; D006899:Mixed Function Oxygenases; D010105:Oxygenases; C021555:methane monooxygenase