Definition of the mitochondrial proteome by measurement of molecular masses of membrane proteins.
The covalent structure of a protein is incompletely defined by its gene sequence, and mass spectrometric analysis of the intact protein is needed to detect the presence of any posttranslational modifications. Because most membrane proteins are purified in detergents that are incompatible with mass spectrometric ionization techniques, this essential measurement has not been made on many hydrophobic proteins, and so proteomic data are incomplete. We have extracted membrane proteins from bovine mitochondria and detergent-purified NADH:ubiquinone oxidoreductase (complex I) with organic solvents, fractionated the mixtures by hydrophilic interaction chromatography, and measured the molecular masses of the intact membrane proteins, including those of six subunits of complex I that are encoded in mitochondrial DNA. These measurements resolve long-standing uncertainties about the interpretation of the mitochondrial genome, and they contribute significantly to the definition of the covalent composition of complex I.
Carroll
Joe
J
Medical Research Council Dunn Human Nutrition Unit, Hills Road, Cambridge CB2 2XY, United Kingdom.
Fearnley
Ian M
IM
Walker
John E
JE
D000818:Animals; D002417:Cattle; D002853:Chromatography, Liquid; D057927:Hydrophobic and Hydrophilic Interactions; D013058:Mass Spectrometry; D008565:Membrane Proteins / Q000737:chemistry / Q000378:metabolism*; D008929:Mitochondria, Heart / Q000737:chemistry / Q000378:metabolism*; D021122:Protein Subunits / Q000737:chemistry / Q000378:metabolism; D020543:Proteome / Q000302:isolation & purification / Q000378:metabolism*; D012997:Solvents; D021241:Spectrometry, Mass, Electrospray Ionization
D008565:Membrane Proteins; D021122:Protein Subunits; D020543:Proteome; D012997:Solvents