Identification by bacterial expression of the yeast genomic sequence encoding L-galactono-γ-lactone oxidase, the homologue of L-ascorbic acid-synthesizing enzyme of higher animals

Autor:	Takahiro Ishikawa, Yuriko Ohta, Morimitsu Nishikimi
Rok vydání:	1998
Předmět:	chemistry.chemical_classification Oxidase test Expression vector biology Clinical Biochemistry Cell Biology Flavin group Ascorbic acid biology.organism_classification Biochemistry Saccharomyces Molecular biology Yeast Amino acid Enzyme chemistry Genetics Molecular Biology
Zdroj:	IUBMB Life. 44:907-913
ISSN:	1521-6543
DOI:	10.1080/15216549800201962
Popis:	Baker's yeast possesses L-galactono-γ-lactone oxidase that is similar in enzymic properties to L-gulono-γ-lactone oxidase, the terminal enzyme of L-ascorbic acid biosynthesis in higher animals. Computer-assisted comparison of amino acid sequences revealed the occurrence of a homologue of rat L-gulono-γ-lactone oxidase in the database of the proteins encoded by the Saccharomyces genome. The gene encoding the homologue was amplified by polymerase chain reaction and expressed as a fusion protein with glutathione S-transferase in Escherichia coli cells with expression vector pGEX-5X-3. The expressed protein was found to show L-galactono-γ-lactone oxidase activity and also to have covalently bound flavin, as is the case with purified L-galactono-γ-lactone oxidase. Thus the gene for this enzyme was unequivocally identified.
Databáze:	OpenAIRE
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