Carbohydrate-binding modules influence substrate specificity of an endoglucanase from Clostridium thermocellum

Figures & data

Fig. 1. Isolated cellulase and carbohydrate-binding module genes. Cel5E from C. thermocellum CelH was used as a model enzyme.

Notes: CBM11 from C. thermocellum CelH, CBM3 from C. josui CipA, and CBM6 from C. stercorarium XynA were also isolated and fused to the C-terminal end of CtCel5E (Yagüe, 1990; Kakiuchi, 1998; Sakka, 1993). Dotted lines show isolated region of the genes.

Fig. 2. Purities of recombinant enzymes.

Notes: M, molecular weight marker (LMW Electrophoresis Calibration Kit; GE Healthcare Life Sciences, Buckinghamshire, UK); lane 1, CtCel5E; lane 2, CtCel5E-CjCBM3; lane 3, CtCel5E-CsCBM6; lane 4, CtCel5E-CtCBM11. The gel was stained with Rapid Stain CBB Kit (Nacalai Tesque, Kyoto, Japan).

Fig. 3. Relative activities of CtCel5E with and without CBMs.

Notes: Enzymatic activities of CtCel5E, CtCel5E-CtCBM11, CtCel5E-CjCBM3, and CtCel5E-CsCBM6 were analyzed with (a) 1.5% carboxymethyl cellulose, (b) 0.5% oat-spelt xylan, (c) 0.5% ball-milled cellulose, and (d) 1% Avicel cellulose at 60 °C. The activities were compared with that of CtCel5E. Bars show standard error.

Fig. 4. Catalytic activities of CtCel5Es for milled corn hulls.

Notes: Reducing sugar release of CtCel5E-CtCBM11 (○), CtCel5E-CjCBM3 (□), and CtCel5E-CsCBM6 (△) from 3 mg milled corn hulls were quantitated. Bars show standard error.