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Multivariate modular engineering of the protein secretory pathway for production of heterologous glucose oxidase in Pichia pastoris

Paper ID Volume ID Publish Year Pages File Format Full-Text
16966 42627 2015 10 PDF Available
Title
Multivariate modular engineering of the protein secretory pathway for production of heterologous glucose oxidase in Pichia pastoris
Abstract

•Individual effects of genes involved in folding machinery and stress responses were investigated.•A modular engineering of protein secretory pathway improved GOD production markedly.•The constructed strain S17 gave a 5.11-fold improvement in GOD activity (1972.9 U/mL).•The GOD activity represented the highest yield reported so far.

Limitations in protein production and secretion have been attributed to the inefficient folding rate of overexpressed proteins and the cellular response to the presence of overexpressed proteins in the endoplasmic reticulum (ER). In this study, we improved the yield of glucose oxidase (GOD) by manipulating genes involved in protein folding machinery and abnormal folding stress responses. First, genes with folding and secretion functions were used to modulate the folding rate of GOD in the ER and its secretion level in the cytoplasm. Next, the potential benefits of the ERAD elements were determined. Cellular resistance to ER derived stress was then strengthened by overexpressing the stress response gene GCN4. Furthermore, a module combination strategy, which co-expressed the SEC53, CNE1 and GCN4 genes, was employed to construct the Pichia pastoris strain S17. This increased the yield of GOD to 21.81 g/L, with an activity of 1972.9 U/mL, which were 2.53- and 5.11-fold higher, respectively, than the control strain. The work described here improved GOD production significantly, and the strategies employed in this study provide novel information for the large-scale production of heterologous proteins.

Keywords
Pichia pastoris; Heterologous protein secretion; Glucose oxidase; Modular engineering
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Multivariate modular engineering of the protein secretory pathway for production of heterologous glucose oxidase in Pichia pastoris
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Publisher
Database: Elsevier - ScienceDirect
Journal: Enzyme and Microbial Technology - Volume 68, January 2015, Pages 33–42
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