Engineering a family 9 processive endoglucanase from Paenibacillus barcinonensis displaying a novel architecture |
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Authors: | Alina Iulia Chiriac Edith Marleny Cadena Teresa Vidal Antonio L Torres Pilar Diaz F I Javier Pastor |
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Institution: | 1. Department of Microbiology, Faculty of Biology, University of Barcelona, Av. Diagonal 645, 08028, Barcelona, Spain 2. Department of Textile and Paper Engineering, ETSEIAT Terrassa, Polytechnic University of Catalonia, Colón 11, 08222 Terrassa, Barcelona, Spain
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Abstract: | Cel9B from Paenibacillus barcinonensis is a modular endoglucanase with a novel molecular architecture among family 9 enzymes that comprises a catalytic domain (GH9),
a family 3c cellulose-binding domain (CBM3c), a fibronectin III-like domain repeat (Fn31,2), and a C-terminal family 3b cellulose-binding domain (CBM3b). A series of truncated derivatives of endoglucanase Cel9B have
been constructed and characterized. Deletion of CBM3c produced a notable reduction in hydrolytic activity, while it did not
affect the cellulose-binding properties as CBM3c did not show the ability to bind to cellulose. On the contrary, CBM3b exhibited
binding to cellulose. The truncated forms devoid of CBM3b lost cellulose-binding ability and showed a reduced activity on
crystalline cellulose, although activity on amorphous celluloses was not affected. Endoglucanase Cel9B produced only a small
ratio of insoluble products from filter paper, while most of the reducing ends produced by the enzyme were released as soluble
sugars (91%), indicating that it is a processive enzyme. Processivity of Cel9B resides in traits contained in the tandem of
domains GH9–CBM3c, although the slightly reduced processivity of truncated form GH9–CBM3c suggests a minor contribution of
domains Fn31,2 or CBM3b, not contained in it, on processivity of endoglucanase Cel9B. |
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