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N-glycosylation enhances functional and structural stability of recombinant β-glucuronidase expressed in Pichia pastoris

Paper ID Volume ID Publish Year Pages File Format Full-Text
23563 43452 2013 7 PDF Available
Title
N-glycosylation enhances functional and structural stability of recombinant β-glucuronidase expressed in Pichia pastoris
Abstract

Recombinant β-glucuronidase (GUS) expressed in Pichia pastoris GS115 is an important glycoprotein, encoded by a gene with four potential N-glycosylation sites. To investigate the impact of N-linked carbohydrate moieties on the stability of recombinant GUS, it was deglycosylated by peptide-N-glycosidase F (PNGase-F) under native conditions. The enzymatic activities of the glycosylated and deglycosylated GUS were compared under various conditions such as temperature, pH, organic solvents, detergents and chaotropic agent. The results demonstrated that the glycosylated GUS retained greater fraction of maximum enzymatic activity against various types of denaturants compared with the deglycosylated. The conformational stabilities of both GUS were analyzed by monitoring the unfolding equilibrium by using the denaturant guanidinium chloride (dn-HCl). The glycosylated GUS displayed a significant increase in its conformational stability than the deglycosylated counterpart. These results affirmed the key role of N-glycosylation on the structural and functional stability of β-glucuronidase and could have potential applications in the functional enhancement of industrial enzymes.

► We report an effective enzymatic deglycosylation for β-glucuronidase (GUS). ► Removal of N-glycosylation reduce the stability of GUS against various denaturants. ► A significant reduction in conformational stability of GUS after deglycosylation. ► These interesting findings are very important to the function enhancement of GUS.

Keywords
β-Glucuronidase; Pichia pastoris; N-glycosylation; Stability; Unfolding equilibrium
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N-glycosylation enhances functional and structural stability of recombinant β-glucuronidase expressed in Pichia pastoris
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Publisher
Database: Elsevier - ScienceDirect
Journal: Journal of Biotechnology - Volume 164, Issue 1, 10 March 2013, Pages 75–81
Authors
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Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering
Get Full-Text Now
Don't Miss Today's Special Offer
Price was $35.95
You save - $31
Price after discount Only $4.95
100% Money Back Guarantee
Full-text PDF Download
Online Support
Any Questions? feel free to contact us