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Heterologous expression of codon optimized Trichoderma reesei Cel6A in Pichia pastoris

Paper ID Volume ID Publish Year Pages File Format Full-Text
16766 42610 2016 10 PDF Available
Title
Heterologous expression of codon optimized Trichoderma reesei Cel6A in Pichia pastoris
Abstract

•Cel6A expressed heterologously in P. pastoris achieved at 1100 U/L with 2 g/L of protein content.•The AOX1 promotor is competitive to the GAP in Cel6A enzyme property and expression.•More efforts should be addressed in a delicate compromise for the high-titer enzyme expression with the high cell biomass.

The Cel6A deficiency has become one of the limiting factors for cellulose saccharification in biochemical conversion of cellulosic biomass to fuels and chemicals. The work attempted to use codon optimization to enhance Trichoderma reesei Cel6A expression in Pichia pastoris. Two recombinants P. pastoris GS115 containing AOX1 and GAP promotors were successfully constructed, respectively. The optimal temperatures and pHs of the expressed Cel6A from two recombinants were consistent with each other, were also in the extremely similar range to that reported on the native Cel6A from T. reesei. Based on the shake flask fermentation, AOX1 promotor enabled the recombinant to produce 265 U/L and 300 mg/L of the Cel6A enzyme, and the GAP promotor resulted in 145 U/L and 200 mg/L. High cell density fed batch (HCDFB) fermentation significantly improved the enzyme titer (1100 U/L) and protein yield (2.0 g/L) for the recombinant with AOX1 promotor. Results have showed that the AOX1 promotor is more suitable than the GAP for the Cel6A expression in P. pastoris. And the HCDFB cultivation is a favorable way to express the Cel6A highly in the methanol inducible yeast.

Keywords
HCDFB, high cell density fed batch; HCD, high cell density; DNS, 3,5-dinitrosalicylic acid; Wd. cbh2, wild cbh2 gene; Opt. cbh2, optimized cbh2 gene; CMC, carboxymethyl cellulose; PASC, phosphoric acid-swollen cellulose; ND, not determined; GAP, glycerald
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Heterologous expression of codon optimized Trichoderma reesei Cel6A in Pichia pastoris
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Publisher
Database: Elsevier - ScienceDirect
Journal: Enzyme and Microbial Technology - Volume 92, October 2016, Pages 107–116
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