کد مقاله کد نشریه سال انتشار مقاله انگلیسی نسخه تمام متن
2817410 1159986 2013 8 صفحه PDF دانلود رایگان
عنوان انگلیسی مقاله ISI
Cloning, characterization and heterelogous expression of the INU1 gene from Cryptococcus aureus HYA
موضوعات مرتبط
علوم زیستی و بیوفناوری بیوشیمی، ژنتیک و زیست شناسی مولکولی ژنتیک
پیش نمایش صفحه اول مقاله
Cloning, characterization and heterelogous expression of the INU1 gene from Cryptococcus aureus HYA
چکیده انگلیسی

The INU1 gene (Accession number: JX073660) encoding exo-inulinase from Cryptococcus aureus HYA was cloned and characterized. The gene had an open reading frame (ORF) of 1653 bp long encoding an inulinase. The coding region of the gene was not interrupted by any intron. It encoded 551 amino acid residues of a protein with a putative signal peptide of 23 amino acids and the calculated molecular mass of 59.5 kDa. The protein sequence deduced from the inulinase structural gene contained the inulinase consensus sequences (WMNDPNGL), (RDP), ECP, FS and Q. It also had two conserved putative N-glycosylation sites. The inulinase from C. aureus HYA was found to be closely related to that from Kluyveromyces marxianus and Pichia guilliermondii. The inulinase gene without the signal sequence was subcloned into pPICZaA expression vector and expressed in Pichia pastoris X-33. The expressed fusion protein was analyzed by SDS-PAGE and western blotting and a specific band with molecular mass of about 60 kDa was found. Enzyme activity assay verified the recombinant protein as an inulinase. A maximum inulinase activity of 16.3 ± 0.24 U/ml was obtained from the culture supernatant of P. pastoris X-33 harboring the inulinase gene. The optimal temperature and pH for action of the enzyme were 50 °C and 5.0, respectively. A large amount of monosaccharides were detected after the hydrolysis of inulin with the purified recombinant inulinase.


► The ORF of the exo-inulinase gene cloned from C. aureus HYA had 1653 bp.
► The cloned gene was expressed in P. pastoris X-33.
► The optimal temperature and pH of the recombinant inulinase were 50 °C and 5.0.
► Only monosaccharide was released from inulin.

ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: Gene - Volume 516, Issue 2, 10 March 2013, Pages 255–262
نویسندگان
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