Mass spectrometric detection of protein, lipid and heme components of cytochrome c oxidase from R-sphaeroides and the stabilization of non-covalent complexes from the enzyme

The cytochrome c oxidase enzyme from Rhodobacter sphaeroides bacteria exists as a complex of four peptide subunits, two hemes and a variety of lipids and metal ions held together by non-covalent forces. Although the native enzyme functions as an associated unit, this complex usually dissociates during matrix-assisted laser desorption/ionization time-of-flight (MALDI-ToF) mass spectrometry analysis. Through the use of matrix additives such as sucrose, the complete complex and partial complexes can be stabilized in the MALDI-ToF experiment. The dissociation of the complex allows the detection of the components of the enzyme. The direct detection of associated lipids from an aqueous solution of the intact enzyme may eliminate the need for enzyme disruption and lipid extraction. The partial dissociation of multi-subunit enzymes in such experiments may allow the determination of subunit-subunit and subunit-lipid interactions.