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Characterization of the family GH54 α-l-arabinofuranosidases in Penicillium funiculosum, including a novel protein bearing a cellulose-binding domain

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Abstract

The soil deuteromycete Penicillium funiculosum is characterized by its remarkable capacity to produce a wide variety of cellulolytic and hemicellulolytic enzymes. In the course of the genome sequencing of this industrial fungus, four different genes encoding glycosyl hydrolase family 54 (GH54)22 α-l-arabinofuranosidases were identified. Three of them termed PfabfB1, PfabfB3, and PfabfB4 were highly similar, encoding proteins of 507, 508, and 505 amino acids, respectively. They exhibited structural features typical of GH54 enzymes, including an N-terminal catalytic domain connected to a C-terminal arabinose-binding domain (ABD). The fourth gene termed PfafbB2 codes for an unusual 400 amino acid length GH54 α-l-arabinofuranosidase, in which the ABD was replaced by a fungal cellulose-binding domain (fCBD). This domain was shown to be functional since it allowed this protein to be retained onto microcrystalline cellulose, and the fusion of this CBD to the C-terminal end of PfAbfB1 allowed this protein to bind to cellulose. Expression analysis of the four PfabfB genes during an industrial-like process fermentation on complex carbohydrates revealed that PfafB2 was expressed more than 20,000-fold, while PfabfB3 and PfabfB4 were increased moderately at the end of the fermentation. In contrast, the transcript levels of PfabfB1 remained unchanged throughout the process. This new type of GH54 α-arabinofuranosidase encoded by PfabfB2 showed enzymatic properties slightly different to those of other GH54 enzymes characterized so far, including a higher thermostability, an optimum pH, and temperature of 2.6 and 50 °C, instead of 3.5 and 60 °C as found for PfAbfB1. Nonetheless, like other GH54 α-arabinofuranosidases, PfAbfB2 was able to release arabinose from various sources of branched arabinoxylan and arabinan.

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Acknowledgments

The technical expertise of Dr. Veronique Anton-Le Berre from the Biochips Platform Toulouse Genopole is gratefully acknowledged for making and using the DNA macroarrays. We also thank Miss Marie-Odile Loret for technical assistance in measuring sugars with HPAEC on Dionex. This work was supported in part by grants from the CINABio-Adisseo company (Antony, France) and from the Region Midi Pyrénées (grant nos. 06001324 and 07006288).

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  1. Olivier Tourrasse

    Present address: METabolic EXplorer, Biopôle Clermont Limagne, 63360, Saint-Beauzire, France

Authors and Affiliations

  1. Université de Toulouse, INSA, UPS, INP, 135 Avenue de Rangueil, 31077, Toulouse, France

    Olivier Guais, Olivier Tourrasse, Marion Dourdoigne, Jean Luc Parrou & Jean Marie Francois

  2. INRA, UMR792 Ingénierie des Systèmes Biologiques et des Procédés, 31400, Toulouse, France

    Jean Marie Francois

  3. CNRS, UMR5504, 31400, Toulouse, France

    Jean Luc Parrou & Jean Marie Francois

  4. Cinabio-Adisseo France S.A.S, 135 Avenue de Rangueil, 31077, Toulouse, France

    Olivier Guais & Olivier Tourrasse

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Guais, O., Tourrasse, O., Dourdoigne, M. et al. Characterization of the family GH54 α-l-arabinofuranosidases in Penicillium funiculosum, including a novel protein bearing a cellulose-binding domain. Appl Microbiol Biotechnol 87, 1007–1021 (2010). https://doi.org/10.1007/s00253-010-2532-4

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