Molecular cloning and over-expression of a fructosyltransferase from Aspergillus niger QU10 / 生物工程学报
Chinese Journal of Biotechnology
;
(12): 512-522, 2015.
Article
in English
| WPRIM
| ID: wpr-240623
ABSTRACT
The main commercial production of fructooligosaccharides (FOS) comes from enzymatic transformation using sucrose as substrate by microbial enzyme fructosyltransferase. A fructosyltransferase genomic DNA was isolated from Aspergillus niger QU10 by PCR. The nucleotide sequence showed a 1 941 bp size, and has been submitted to GenBank (KF699529). The cDNA of the fructosyltransferase, containing an open reading frame of 1 887 bp, was further cloned by RT-PCR. The fructosyltransferase gene from Aspergillus niger was functionally expressed both in Escherichia coli and Pichia pastoris GS 115. The highest activity value for the construction with the α-factor signal peptide reached 431 U/mL after 3 days of incubation. The recombinant enzyme is extensively glycosylated, and the active form is probably represented by a homodimer with an apparent molecular mass of 200 kDa as judged from mobility in seminative PAGE gels. The extracellular recombinant enzyme converted sucrose mostly to FOS, mainly 1-kestose and nystose, liberating glucose. FOS reached a maximal value and represented about 58% of total sugars present in the reaction mixture after 4 h reaction. The results suggest that the availability of recombinant Pichia pastoris as a new source of a FOS-producing enzyme might result of biotechnology interest for industrial application.
Full text:
Available
Index:
WPRIM (Western Pacific)
Main subject:
Pichia
/
Aspergillus niger
/
Sucrose
/
Trisaccharides
/
Glycosylation
/
Fungal Proteins
/
Molecular Sequence Data
/
Base Sequence
/
Cloning, Molecular
/
DNA, Complementary
Language:
English
Journal:
Chinese Journal of Biotechnology
Year:
2015
Type:
Article
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