Cloning, expression, purification and assay of sorbitol dehydrogenase from "Feicheng" peach fruit (Prunus persica)
Braz. arch. biol. technol
;
56(4): 531-539, July-Aug. 2013. ilus, graf
Artigo
em Inglês
| LILACS
| ID: lil-684504
ABSTRACT
A gene encoding NAD+-dependent sorbitol dehydrogenase (SDH) in peach fruit was cloned and expressed in Escherichia coli. Recombinant SDH protein with 6×His-tagged was localized exclusively in the cytoplasmic soluble fraction of E. coli when the strains were grown for 4-5 h at 37 ºC. Highly pure protein was isolated by Ni2+-resin chromatography with 150 mM imidazole in 50 mM Tris, pH 8.0, by elution. In order to ensure that the recombinant SDH could be used for further study, the fluorescence and ultraviolet spectrum of the recombinant SDH were detected. Recombinant SDH was confirmed to be capable of oxidizing sorbitol by enzymatic activity assay. The activity of the recombinant SDH was 2.73 U mg-1min-1, which was similar with that directly extracted from peach fruits. The activities of SDH extracted from the fruits in different periods (30, 60, 90 days after flowing) were 7.75, 5.95, 3.26 U mg-1min-1, respectively.
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Índice:
LILACS (Américas)
Idioma:
Inglês
Revista:
Braz. arch. biol. technol
Assunto da revista:
Biologia
Ano de publicação:
2013
Tipo de documento:
Artigo
País de afiliação:
China
Instituição/País de afiliação:
Shandong Agricultural University/CN
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