title: Purification and properties of a novel broad substrate specific alcohol oxidase from Aspergillus terreus MTCC 6324.
abstract: An alcohol oxidase was isolated from the microsome of n-hexadecane grown Aspergillus terreus and purified by ion exchange chromatography. The oxidase was found to act on short chain-, long chain-, secondary-, and aromatic-alcohol substrates with highest affinity for n-heptanol (K(M)=0.498 mM, K(cat)=2.7x10(2) s(-1)). The native protein molecular mass was 269+/-5 kDa and the subunit molecular masses were 85-, 63-, 43-, 27-, and 13-kDa. The isoelectric point of the proteins was within 8.3-8.5. High aggregating property of the protein was demonstrated by AFM, DLS and TEM analyses. Chemical analysis showed the presence of oleic acid and palmitic acid at a ratio of 2:1 in the purified protein. This lipoidic nature of the protein particles was correlated to the high aggregating property. In this flavoenzyme, flavin was non-covalently but avidly associated. Peptide mass fingerprinting studies showed the presence of two FAD binding domains in 63 kDa protein. Among these two FAD binding domain sequences only the YPVIDHEYDAVVVGAGGAGLR peptide shows 45-50% sequence homology with the reported N-terminal sequences of other known alcohol oxidases. Non-redundant database search of 63- and 43-kDa subunits peptide sequences showed no sequence similarity with the other alcohol oxidase protein reported till now.
authors.affiliation: Department of Biotechnology, Indian Institute of Technology Guwahati, Guwahati-781039, Assam, India.
authors.forename: Adepu Kiran
authors.initials: AK
authors.lastname: Kumar
authors.forename: Pranab
authors.initials: P
authors.lastname: Goswami
mesh_terms: D000429:Alcohol Oxidoreductases / Q000737:chemistry* / Q000302:isolation & purification* / Q000378:metabolism; D000595:Amino Acid Sequence; D001230:Aspergillus / Q000737:chemistry* / Q000201:enzymology; D007700:Kinetics; D008969:Molecular Sequence Data; D010084:Oxidation-Reduction; D011487:Protein Conformation; D021241:Spectrometry, Mass, Electrospray Ionization; D013379:Substrate Specificity
chemical_list: D000429:Alcohol Oxidoreductases; C019643:alcohol oxidase