Engineering a family 9 processive endoglucanase from Paenibacillus barcinonensis displaying a novel architecture.
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 (Fn3(1,2)), 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 Fn3(1,2) or CBM3b, not contained in it, on processivity of endoglucanase Cel9B.
Chiriac
Alina Iulia
AI
Department of Microbiology, Faculty of Biology, University of Barcelona, Av. Diagonal 645, Barcelona, Spain.
Cadena
Edith Marleny
EM
Vidal
Teresa
T
Torres
Antonio L
AL
Diaz
Pilar
P
Pastor
F I Javier
FI
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D002480:Cellulase / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D002482:Cellulose / Q000378:metabolism; D008969:Molecular Sequence Data; D056507:Paenibacillus / Q000201:enzymology* / Q000235:genetics*; D011485:Protein Binding; D015202:Protein Engineering; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment; D017384:Sequence Deletion
D001426:Bacterial Proteins; D011994:Recombinant Proteins; D002482:Cellulose; D002480:Cellulase