Expression inEscherichia coliof an Unnamed Protein Gene fromAspergillus oryzaeRIB40 and Cofactor Analyses of the Gene Product as Formate Oxidase
| Autor: | Hiroaki Nose, Masaya Oki, Kimio Isa, Yoshifumi Maeda, Yutaka Fujii, Akihiko Sakurai, Hiroyuki Uchida, Daiju Doubayashi |
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| Rok vydání: | 2009 |
| Předmět: |
Formates
Aspergillus oryzae Coenzymes Glucose-methanol-choline oxidoreductase family Gene Expression medicine.disease_cause Applied Microbiology and Biotechnology Biochemistry Cofactor Substrate Specificity Analytical Chemistry chemistry.chemical_compound Affinity chromatography Escherichia coli medicine Formate Molecular Biology chemistry.chemical_classification Oxidase test biology Protein Stability Organic Chemistry Temperature Hydrogen Peroxide General Medicine Hydrogen-Ion Concentration biology.organism_classification Molecular Weight Enzyme chemistry Biocatalysis Flavin-Adenine Dinucleotide biology.protein Spectrophotometry Ultraviolet Oxidoreductases Oxidation-Reduction Biotechnology |
| Zdroj: | Bioscience, Biotechnology, and Biochemistry. 73:2645-2649 |
| ISSN: | 1347-6947 0916-8451 |
| DOI: | 10.1271/bbb.90497 |
| Popis: | An unnamed protein of Aspergillus oryzae RIB40 (accession no. XP_001727378), the amino acid sequence of which shows high similarity to those of formate oxidase isoforms produced by Debaryomyces vanjiriae MH201, was produced in Escherichia coli in C-His(6)-tagged form. The gene product, purified by affinity column chromatography, catalyzed the oxidation of formate to yield hydrogen peroxide but showed no evidence of activity on the other substrates tested. The K(m) and V(max) values at 30 degrees C at pH 4.5 were 7.9 mM and 26.3 micromole/min mg respectively. The purified enzyme showed UV-visible spectra atypical of ordinary flavoproteins. The UV-visible spectra of the enzyme and the UV-visible spectra, fluorescence spectra, and mass spectrometry of the extract obtained by boiling the purified enzyme suggested that the enzyme has a non-covalently bound FAD analog, which is expected to be 8-formyl-FAD. |
| Databáze: | OpenAIRE |
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