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Sammanfattning

A novel alcohol oxidase (AOX) has been purified from mycelial pellets of the wood degrading basidiomycete Gloeophyllum trabeum and characterized as a homooctameric nonglycosylated protein with native and subunit molecular masses of 628 and 72.4 kDa, containing noncovalently bonded flavin adenine dinucleotide. The isolated AOX cDNA contained an open reading frame of 1953 bp translating into a polypeptide of 651 amino acids displaying 51-53% identity with other published fungal AOX amino acid sequences. The enzyme catalyzed oxidation of short chain primary aliphatic alcohols with preference for methanol (Km 2.3 mM, kcat 15.6 s(-1)). Using polyclonal antibodies and immunofluoresence staining, AOX was localized on liquid culture hyphae and extracellular slime, in sections from degraded wood and on cotton fibres. Transmission electron microscopy immunogold labeling localized the enzyme in the hyphal periplasmic space and wall, and on extracellular tripartite membranes and slime, while there was no labeling of hyphal peroxisomes. AOX was further shown associated with membraneous/slime structures secreted by hyphae in wood fibre lumina and within secondary cell walls of degraded wood fibres. The differences in AOX targeting compared with the known yeast peroxisomal localization were traced to a unique C-terminal sequence of the G. trabeum oxidase, apparently responsible for the protein's different translocation. The extracellular distribution, and the enzyme's abundance and preference for methanol, potentially available from demethylation of lignin, all point to a possible role for AOX as a major source of H2O2, a component of Fenton's reagent implicated in the generally accepted mechanisms for brown rot through production of highly destructive hydroxyl radicals

Publicerad i

Applied and Environmental Microbiology
2007, volym: 73, nummer: 19, sidor: AEM.00977-07
Utgivare: American Society for Microbiology

SLU författare

  • Daniel, Geoffrey

    • Institutionen för skogens produkter, Sveriges lantbruksuniversitet

  • Stålhandske, Lada

    • Institutionen för skogens produkter, Sveriges lantbruksuniversitet

UKÄ forskningsämne

Skogsvetenskap

Publikationens identifierare

  • DOI: https://doi.org/10.1128/AEM.00977-07

Permanent länk till denna sida (URI)

https://res.slu.se/id/publ/14934