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High-Level Heterologous Expression of Endo-1,4-β-Xylanase from Penicillium citrinum in Pichia pastoris X-33 Directed through Codon Optimization and Optimized Expression

Abstract : Most common industrial xylanases are produced from filamentous fungi. In this study, the codon-optimized xynA gene encoding xylanase A from the fungus Penicilium citrinum was successfully synthesized and expressed in the yeast Pichia pastoris. The levels of secreted enzyme activity under the control of glyceraldehyde-3-phosphate dehydrogenase (P GAP) and alcohol oxidase 1 (P AOX1) promoters were compared. The Pc Xyn11A was produced as a soluble protein and the total xylanase activity under the control of P GAP and P AOX1 was 34-and 193-fold, respectively, higher than that produced by the native strain of P. citrinum. The Pc Xyn11A produced under the control of the P AOX1 reached a maximum activity of 676 U/mL when induced with 1% (v/v) methanol every 24 h for 5 days. The xylanase was purified by ion exchange chromatography and then characterized. The enzyme was optimally active at 55 • C and pH 5.0 but stable over a broad pH range (3.0-9.0), retaining more than 80% of the original activity after 24 h or after pre-incubation at 40 • C for 1 h. With birchwood xylan as a substrate, Pc Xyn11A showed a K m(app) of 2.8 mg/mL, and a k cat of 243 s −1. The high level of secretion of Pc Xyn11A and its stability over a wide range of pH and moderate temperatures could make it useful for a variety of biotechnological applications.
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Chanika Ouephanit, Nassapat Boonvitthya, Sophie Bozonnet, Warawut Chulalaksananukul. High-Level Heterologous Expression of Endo-1,4-β-Xylanase from Penicillium citrinum in Pichia pastoris X-33 Directed through Codon Optimization and Optimized Expression. Molecules, MDPI, 2019, 24 (19), ⟨10.3390/molecules24193515⟩. ⟨hal-02966690⟩

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