Homologous expression of Phanerochaete chrysosporium manganese peroxidase, using bialaphos resistance as a dominant selectable marker.
Manganese peroxidase (MnP) is a major extracellular component of the lignin-degrading system of the white-rot fungus, Phanerochaete chrysosporium. Homologous expression of recombinant MnP isozyme 1 (rMnP1) in P. chrysosporium was achieved using a novel transformation system for this fungus, which utilizes the Streptomyces hygroscopicus bialaphos-resistant gene, bar, as the selectable marker. The transformation frequency for this system is approximately 100 bialaphos-resistant transformants per microgram of plasmid DNA. Transformed strains all contain plasmid DNA, ectopically integrated into the fungal genome. Using this transformation system, the promoter region of the P. chrysosporium translation elongation factor gene was used to drive expression of mnp1, encoding MnP1, in primary metabolic cultures of P. chrysosporium, where endogenous MnP was not expressed. Approximately 2-3 mg of active recombinant MnP1 per liter of extracellular medium was produced in agitated cultures of transformants.
Ma
Biao
B
Department of Biochemistry and Molecular Biology, OGI School of Science and Engineering, Oregon Health & Science University, 20000 N.W. Walker Road, Beaverton, OR 97006-8921, USA.
Mayfield
Mary B
MB
Gold
Michael H
MH
D000613:Aminobutyrates / Q000494:pharmacology; D000935:Antifungal Agents / Q000494:pharmacology; D001483:Base Sequence; D004351:Drug Resistance / Q000235:genetics; D005786:Gene Expression Regulation; D005799:Genes, Dominant; D005819:Genetic Markers; D005822:Genetic Vectors; D008969:Molecular Sequence Data; D009943:Organophosphorus Compounds / Q000494:pharmacology*; D010544:Peroxidases / Q000235:genetics* / Q000378:metabolism; D020075:Phanerochaete / Q000201:enzymology* / Q000235:genetics*; D010957:Plasmids; D011401:Promoter Regions, Genetic; D011994:Recombinant Proteins / Q000187:drug effects / Q000235:genetics; D014170:Transformation, Genetic
D000613:Aminobutyrates; D000935:Antifungal Agents; D005819:Genetic Markers; D009943:Organophosphorus Compounds; D011994:Recombinant Proteins; C003122:bialaphos; C003121:phosphinothricin; D010544:Peroxidases; C051129:manganese peroxidase