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Screening of signal sequences for extracellular production of Aspergillus niger xylanase in Pichia pastoris

Paper ID Volume ID Publish Year Pages File Format Full-Text
3093 150 2014 6 PDF Available
Title
Screening of signal sequences for extracellular production of Aspergillus niger xylanase in Pichia pastoris
Abstract

•The xylanase B gene was expressed with four different secretion signals in P. pastoris.•Highest xylanase activity among single-copy clones was obtained with PIR1 secretion sequence.•The xylanase was purified with a single step Ni-NTA purification method.•The optimum temperature and pH for recombinant xylanase was 40–50 °C and pH 5, respectively.•Specific activity of purified xylanase was 13,271 U/mg.

In this study four secretion signal sequences were evaluated for extracellular production of Aspergillus niger xylanase (XylB) in Pichia pastoris. These sequences were namely; Saccharomyces cerevisiae α-mating factor (α-MF), Human Serum Albumin (HSA), P. pastoris Acid Phosphatase (PHO1) and P. pastoris Protein with Internal Repeats (PIR1) secretion signals. The codon optimized XylB gene fused to secretion sequences was expressed under the AOX1 promoter of P. pastoris in KM71H strain. Xylanase enzyme activities for single-copy clones with α-MF, HSA, PHO1 and PIR1 as secretion sequence were 170, 123, 139 and 250 U/mL, respectively, after 48 h induction period. P. pastoris PIR1 leader sequence resulted highest xylanase enzyme activity at shake-flask scale among the tested secretion signal sequences. The xylanase was purified with a single step Ni-NTA purification method. The optimum temperature and pH for recombinant xylanase was 40–50 °C and pH 5, respectively. Additionally, xylanase activity in purified enzyme was stable over a wide pH range (pH 3–7) and temperature of up to 50 °C. Specific activity of purified xylanase was 13,271 U/mg.

Keywords
Yeast; Enzyme production; Purification; Optimization; Pichia pastoris; Xylanase
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Screening of signal sequences for extracellular production of Aspergillus niger xylanase in Pichia pastoris
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Publisher
Database: Elsevier - ScienceDirect
Journal: Biochemical Engineering Journal - Volume 92, 15 November 2014, Pages 16–21
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