Biochemical characterization of MmoS, a sensor protein involved in copper-dependent regulation of soluble methane monooxygenase.
Methane monooxygenase (MMO) enzymes catalyze the oxidation of methane to methanol in methanotrophic bacteria. Several strains of methanotrophs, including Methylococcus capsulatus (Bath), express a membrane-bound or particulate MMO (pMMO) at high copper-to-biomass ratios and a soluble MMO (sMMO) form when copper is limited. The mechanism of this "copper switch" is not understood. The mmoS gene, located downstream of the sMMO operon, encodes a sensor protein that is part of a two-component signaling system and has been proposed to play a role in the copper switch. MmoS from M. capsulatus (Bath) has been cloned, expressed, and purified. The purified protein is a tetramer of molecular mass 480 kDa. Optical spectra indicate that MmoS contains a flavin cofactor, identified as flavin adenine dinucleotide (FAD) by fluorescence spectroscopy and chromatographic analysis. The redox potential of the MmoS-bound FAD, which binds within the N-terminal PAS-PAC domains, is -290 +/- 2 mV at pH 8.0 and 25 degrees C. Despite extensive efforts, MmoS could not be loaded with Cu(I) or Cu(II), indicating that MmoS does not sense copper directly. These data suggest that MmoS functions as a redox sensor and provide new insight into the copper-mediated regulation of sMMO expression.
Ukaegbu
Uchechi E
UE
Department of Biochemistry, Molecular Biology, and Cell Biology and Department of Chemistry, Northwestern University, Evanston, Illinois 60208, USA.
Henery
Shannon
S
Rosenzweig
Amy C
AC
D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D002853:Chromatography, Liquid / Q000379:methods; D003300:Copper / Q000737:chemistry* / Q000378:metabolism / Q000494:pharmacology; D005182:Flavin-Adenine Dinucleotide / Q000737:chemistry* / Q000378:metabolism; D015964:Gene Expression Regulation, Bacterial / Q000187:drug effects / Q000502:physiology; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects / Q000502:physiology; D020619:Methylococcus capsulatus / Q000201:enzymology* / Q000235:genetics; D009876:Operon / Q000235:genetics; D010105:Oxygenases / Q000096:biosynthesis / Q000737:chemistry* / Q000235:genetics; D011485:Protein Binding; D020836:Protein Structure, Quaternary; D017434:Protein Structure, Tertiary; D013050:Spectrometry, Fluorescence / Q000379:methods
D001426:Bacterial Proteins; D005182:Flavin-Adenine Dinucleotide; D003300:Copper; D010105:Oxygenases; C021555:methane monooxygenase