Purified particulate methane monooxygenase from Methylococcus capsulatus (Bath) is a dimer with both mononuclear copper and a copper-containing cluster.
Particulate methane monooxygenase (pMMO) is a membrane-bound enzyme that catalyzes the oxidation of methane to methanol in methanotropic bacteria. Understanding how this enzyme hydroxylates methane at ambient temperature and pressure is of fundamental chemical and potential commercial importance. Difficulties in solubilizing and purifying active pMMO have led to conflicting reports regarding its biochemical and biophysical properties, however. We have purified pMMO from Methylococcus capsulatus (Bath) and detected activity. The purified enzyme has a molecular mass of approximately 200 kDa, probably corresponding to an alpha(2)beta(2)gamma(2) polypeptide arrangement. Each 200-kDa pMMO complex contains 4.8 +/- 0.8 copper ions and 1.5 +/- 0.7 iron ions. Electron paramagnetic resonance spectroscopic parameters corresponding to 40-60% of the total copper are consistent with the presence of a mononuclear type 2 copper site. X-ray absorption near edge spectra indicate that purified pMMO is a mixture of Cu(I) and Cu(II) oxidation states. Finally, extended x-ray absorption fine structure data are best fit with oxygennitrogen ligands and a 2.57-A Cu-Cu interaction, providing direct evidence for a copper-containing cluster in pMMO.
Lieberman
Raquel L
RL
Department of Biochemistry, Northwestern University, Evanston, IL 60208, USA.
Shrestha
Deepak B
DB
Doan
Peter E
PE
Hoffman
Brian M
BM
Stemmler
Timothy L
TL
Rosenzweig
Amy C
AC
D002458:Cell Fractionation / Q000379:methods; D002462:Cell Membrane / Q000201:enzymology; D002942:Circular Dichroism; D003300:Copper / Q000032:analysis*; D019281:Dimerization; D004578:Electron Spin Resonance Spectroscopy; D007700:Kinetics; D008667:Metalloproteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D020619:Methylococcus capsulatus / Q000201:enzymology*; D008970:Molecular Weight; D010105:Oxygenases / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*; D011487:Protein Conformation
D008667:Metalloproteins; D003300:Copper; D010105:Oxygenases; C021555:methane monooxygenase