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.
Visser
Nina V
NV
Eukaryotic Microbiology, Groningen Biomolecular Sciences and Biotechnology Institute, University of Groningen, P.O. Box 14, 9750AA, Haren, The Netherlands.
Wang
Dongyuan
D
Stanley
Will A
WA
Groves
Matthew R
MR
Wilmanns
Matthias
M
Veenhuis
Marten
M
van der Klei
Ida J
IJ
D000429:Alcohol Oxidoreductases / Q000737:chemistry*; D019281:Dimerization; D004121:Dimethyl Sulfoxide / Q000737:chemistry*; D004789:Enzyme Activation; D004795:Enzyme Stability; D012995:Solubility
D000429:Alcohol Oxidoreductases; C019643:alcohol oxidase; D004121:Dimethyl Sulfoxide