Production by a baculovirus expression system of the APO-protein of L-gulono-gamma-lactone oxidase, a flavoenzyme possessing a covalently-bound FAD.
L-Gulono-gamma-lactone oxidase, an enzyme functioning in L-ascorbic acid biosynthesis in higher animals, possesses a covalently-bound FAD as the prosthetic group. Catalytically-active enzyme was expressed in silkworm cells by a recombinant baculovirus encoding rat L-gulono-gamma-lactone oxidase. When recombinant enzyme was expressed under riboflavin-deficient conditions, most of it was found to be the apoprotein, as evidenced by an increase in enzymic activity upon addition of FAD to the assay mixture. Interestingly, the observed enzymic activity is thought to have been provoked by a noncovalent interaction between FAD and the apoprotein, since the covalent attachment of FAD was not demonstrated by a fluorometric gel-scanning experiment.
Institute of Applied Biochemistry, Yagi Memorial Park, Gifu, Japan.
M
M
Nishikimi
J
J
Kobayashi
K
K
Yagi
D000818:Animals; D001059:Apoproteins / Q000378:metabolism*; D016367:Baculoviridae / Q000378:metabolism*; D015153:Blotting, Western; D012831:Bombyx; D004591:Electrophoresis, Polyacrylamide Gel; D005182:Flavin-Adenine Dinucleotide / Q000378:metabolism*; D015967:Gene Expression Regulation, Viral; D015151:Immunoblotting; D050544:L-Gulonolactone Oxidase; D008297:Male; D008862:Microsomes, Liver / Q000201:enzymology; D051381:Rats; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000378:metabolism; D013401:Sugar Alcohol Dehydrogenases / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics
D001059:Apoproteins; D011994:Recombinant Proteins; D005182:Flavin-Adenine Dinucleotide; D013401:Sugar Alcohol Dehydrogenases; D050544:L-Gulonolactone Oxidase