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Effect of glycosylation on biochemical characterization of recombinant phytase expressed in Pichia pastoris

Paper ID Volume ID Publish Year Pages File Format Full-Text
18269 42716 2008 6 PDF Available
Title
Effect of glycosylation on biochemical characterization of recombinant phytase expressed in Pichia pastoris
Abstract

Recombinant phytase expressed in Pichia pastoris FPHY34 is a typical glycoprotein, whose gene possesses 10 potential N-glycosylation sites without O-glycosylation. Effects of glycosylation on phytase's biochemical characterization were investigated. Secreted phytase's micro-heterogenious glycosylation phenomenon was observed and verified by SDS–PAGE, N-terminal amino acid sequencing and mass spectrum analyses. Deglycosylation of recombinant phytase reduced the molecular weight from 83 to 63 kDa, which indicates the presence of total carbohydrate content of approximately 24.1%. According to image analysis of 2-dimension gel electrophoresis, the range of isoeclectric point of phytase covers nearly 1 pH value rather than a specific point, which might be due to heavy and complex glycosylation. The phytase expressed by P. pastoris still retained 40% and 30% activities at 80 and 90 °C for 10 min, respectively. However, upon deglycosylation in vitro by peptide-N-glycohydrolase F (PNGase F), thermostability of deglycosylated phytase significantly declined after 10 min treatment at 40 and 50 °C. Optimum pH of phytase tested was shifted from pH 5.0 to 2.5 by deglycosylation, although there exhibit bi-humps pH optima at both before and after PNGase F digestion cases.

Keywords
Phytase; Glycosylation; Biochemical characterization; Thermostability; Micro-heterogeneity
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Effect of glycosylation on biochemical characterization of recombinant phytase expressed in Pichia pastoris
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Publisher
Database: Elsevier - ScienceDirect
Journal: Enzyme and Microbial Technology - Volume 42, Issue 4, 4 March 2008, Pages 340–345
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