Dissociation and reconstitution studies of a broad substrate specific multimeric alcohol oxidase protein produced by Aspergillus terreus.
A multimeric alcohol oxidase from Aspergillus terreus was dissociated and simultaneously deflavinated into catalytically inactive FAD-free subunits when incubated with 0.74 M beta-mercaptoethanol (beta-ME) for 8 h at 4 degrees C. This dissociation process had traversed through two FAD-associated intermediate proteins, between these one of them showed the enzyme activity. On removal of beta-ME, the multimeric apoprotein was regenerated, which was, however, catalytically inactive. Reactivation of the FAD supplemented apoprotein was accomplished only after incubating with the substrate. This catalytic reactivation was a slow process as evident from the prolonged FAD emission quenching. The dissociation and re-association phenomena were demonstrated by using dynamic light scattering, size exclusion chromatographic, confocal laser scan microscopic and native PAGE analyses. The solvent effect caused by the high concentration of beta-ME is attributed to the observed dissociation and linked deflavination of these multimeric alcohol oxidase protein particles.
Kumar
Adepu Kiran
AK
Department of Biotechnology, Indian Institute of Technology Guwahati, Guwahati 781039, India.
Goswami
Pranab
P
D000429:Alcohol Oxidoreductases / Q000737:chemistry*; D001059:Apoproteins / Q000737:chemistry; D001230:Aspergillus / Q000201:enzymology* / Q000378:metabolism; D002384:Catalysis; D002850:Chromatography, Gel; D007700:Kinetics; D013379:Substrate Specificity
D001059:Apoproteins; D000429:Alcohol Oxidoreductases; C019643:alcohol oxidase