Dissociation and Reconstitution Studies of a Broad Substrate Specific Multimeric Alcohol Oxidase Protein Produced by Aspergillus terreus

Authors: Kumar, Adepu Kiran; Goswami, Pranab

Source: The Journal of Biochemistry, Volume 145, Number 2, 8 February 2009 , pp. 259-265(7)

Buy & download fulltext article:

Price: $42.29 plus tax (Refund Policy)

Abstract:

A multimeric alcohol oxidase from Aspergillus terreus was dissociated and simultaneously deflavinated into catalytically inactive FAD-free subunits when incubated with 0.74 M β-mercaptoethanol (β-ME) for 8 h at 4°C. This dissociation process had traversed through two FAD-associated intermediate proteins, between these one of them showed the enzyme activity. On removal of β-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 β-ME is attributed to the observed dissociation and linked deflavination of these multimeric alcohol oxidase protein particles.

Keywords: alcohol oxidase; β-mercaptoethanol; deflavination; dissociation; reactivation

Document Type: Regular paper

DOI: http://dx.doi.org/10.1093/jb/mvn163

Publication date: 2009-02-08