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Improved production of two expansin-like proteins in Pichia pastoris and investigation of their functional properties

Paper ID Volume ID Publish Year Pages File Format Full-Text
3237 160 2014 12 PDF Available
Title
Improved production of two expansin-like proteins in Pichia pastoris and investigation of their functional properties
Abstract

•Yield of TrSwo1/BsEXLX1 is significantly improved compared with previous reports.•We firstly reveal that the synergism of TrSwol is greater than that of BsEXLX1.•The unreported functional properties of BsEXLX1/TrSwol are firstly demonstrated.•BsEXLX1/TrSwol exhibited relatively high thermal stability at temperature 50–90 °C.•BsEXLX1/TrSwol showed resistance to denaturation by urea and SDS.

The genes encoding expanin-like proteins from Trichoderma reesei (TrSwo1) and Bacillus subtilis (BsEXLX1) were successfully expressed in Pichia pastoris. The yields of two recombinant proteins were significantly improved by the use of PMSF (phenylmethylsulfonyl fluoride) and a commercial protease inhibitor cocktail. Under the optimum culture conditions, the highest TrSwo1/BsEXLX1 expression level reached was approximately 120/860 mg l−1, which was almost 2.4/86-fold as much as the highest expression level in other host cells. Purified BsEXLX1/TrSwo1 displayed synergism in cellulose hydrolysis with endoglucanase, and the maximum amount of reducing sugars released was almost 2.0/2.5-fold as high as those in reaction mixtures without expansin-like proteins. The synergistic effect reached the maximum level when 1 mg of target protein per g of filter paper was loaded. Both proteins exhibited relatively high thermal stability at temperatures of 50, 70 and 90 °C, and retained more than 45% residual activities after 1 h of pre-incubation at 100 °C, suggesting remarkable heat tolerance. They also showed resistance to denaturation by urea and SDS. Under several enzymatic hydrolysis conditions, the synergistic activity of TrSwo1 was higher than that of BsEXLX1, indicating stronger disrupting activity of TrSwo1 on cellulose than BsEXLX1. This is the first study to report high-efficient expression and unreported properties of BsEXLX1/TrSwo1.

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Keywords
CUTME, cOmplete ULTRA Tablets; PVDF, polyvinylidene difluoride membrane; IOD, integrated optical densityCellulose; Bioconversion; Protein; Pichia pastoris; Optimization; Functional properties
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Improved production of two expansin-like proteins in Pichia pastoris and investigation of their functional properties
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Publisher
Database: Elsevier - ScienceDirect
Journal: Biochemical Engineering Journal - Volume 84, 15 March 2014, Pages 16–27
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