Octameric alcohol oxidase dissociates into stable, soluble monomers upon incubation with dimethylsulfoxide.

Alcohol oxidase (AO) is a peroxisomal, homo-octameric flavoenzyme, which catalyzes methanol oxidation in methylotrophic yeast. Here, we report on the generation of soluble, FAD-lacking AO monomers. Using steady-state fluorescence, fluorescence correlation spectroscopy, circular dichroism and static light scattering approaches, we demonstrate that FAD-lacking AO monomers are formed upon incubation of purified, native octameric AO in a solution containing 50% dimethylsulfoxide (DMSO). Upon removal of DMSO the protein remained monomeric and soluble and did not contain FAD. Binding experiments revealed that the AO monomers bind to purified pyruvate carboxylase, a protein that plays a role in the formation of enzymatically active AO octamers in vivo.