Recombinant Expression of Trichoderma reesei Cel61A in Pichia pastoris: Optimizing Yield and N-terminal Processing

Magali Tanghe, Barbara Danneels, Andrea Camattari, Anton Glieder, Isabel Vandenberghe, Bart Devreese, Ingeborg Stals, Tom Desmet

Publikation: Beitrag in einer FachzeitschriftArtikelForschungBegutachtung

Abstract

The auxiliary activity family 9 (AA9, formerly GH61) harbors a recently discovered group of oxidative enzymes that boost cellulose degradation. Indeed, these lytic polysaccharide monooxygenases (LPMOs) are able to disrupt the crystalline structure of cellulose, thereby facilitating the work of hydrolytic enzymes involved in biomass degradation. Since these enzymes require an N-terminal histidine residue for activity, their recombinant production as secreted protein is not straightforward. We here report the expression optimization of Trichoderma reesei Cel61A (TrCel61A) in the host Pichia pastoris. The use of the native TrCel61A secretion signal instead of the alpha-mating factor from Saccharomyces cerevisiae was found to be crucial, not only to obtain high protein yields (>400 mg/L during fermentation) but also to enable the correct processing of the N-terminus. Furthermore, the LPMO activity of the enzyme is demonstrated here for the first time, based on its degradation profile of a cellulosic substrate.

Originalspracheenglisch
Seiten (von - bis)1010-7
Seitenumfang8
FachzeitschriftMolecular biotechnology
Jahrgang57
Ausgabenummer11-12
DOIs
PublikationsstatusVeröffentlicht - Dez 2015

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Trichoderma
Pichia
Enzymes
Mating Factor
Polysaccharides
Processing
Mixed Function Oxygenases
Cellulose
Degradation
Proteins
Ports and harbors
Histidine
Yeast
Biomass
Fermentation
Crystalline materials
Substrates

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    Recombinant Expression of Trichoderma reesei Cel61A in Pichia pastoris : Optimizing Yield and N-terminal Processing. / Tanghe, Magali; Danneels, Barbara; Camattari, Andrea; Glieder, Anton; Vandenberghe, Isabel; Devreese, Bart; Stals, Ingeborg; Desmet, Tom.

    in: Molecular biotechnology, Jahrgang 57, Nr. 11-12, 12.2015, S. 1010-7.

    Publikation: Beitrag in einer FachzeitschriftArtikelForschungBegutachtung

    Tanghe, M, Danneels, B, Camattari, A, Glieder, A, Vandenberghe, I, Devreese, B, Stals, I & Desmet, T 2015, 'Recombinant Expression of Trichoderma reesei Cel61A in Pichia pastoris: Optimizing Yield and N-terminal Processing' Molecular biotechnology, Jg. 57, Nr. 11-12, S. 1010-7. https://doi.org/10.1007/s12033-015-9887-9
    Tanghe, Magali ; Danneels, Barbara ; Camattari, Andrea ; Glieder, Anton ; Vandenberghe, Isabel ; Devreese, Bart ; Stals, Ingeborg ; Desmet, Tom. / Recombinant Expression of Trichoderma reesei Cel61A in Pichia pastoris : Optimizing Yield and N-terminal Processing. in: Molecular biotechnology. 2015 ; Jahrgang 57, Nr. 11-12. S. 1010-7.
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    KW - Molecular Sequence Data

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