Creation of a cellooligosaccharide-assimilating Escherichia coli strain by displaying active beta-glucosidase on the cell surface via a novel anchor protein

Abstract

We demonstrated direct assimilation of cellooligosaccharide using Escherichia coli displaying beta-glucosidase (BGL). BGL from Thermobifida fusca YX (Tfu0937) was displayed on the E. coli cell surface using a novel anchor protein named Blc. This strain was grown successfully on 0.2% cellobiose, and the optical density at 600 nm (OD(600)) was 1.05 after 20 h.

Fig. 1.

BGL activity on the E. coli cell surface or in culture supernatant after 24 h of cultivation in LB medium. Gray bars show BGL activity on the cell surface (U/OD₆₀₀/ml; left axis), and white bars show BGL activity in the culture supernatant (U/ml; right axis). (A and B) Each BGL was displayed on the E. coli cell surface using the PgsA anchor protein. (A) E. coli BW25113; (B) E. coli JCM20137. (C and D) Tfu0937 was displayed using various anchor proteins. In the case of HdeD-Tfu0937, Blc-Tfu0937, and Slp-Tfu0937, the C terminus of each anchor protein was fused to the N terminus of Tfu0937. In the case of Tfu0937-HdeD, Tfu0937-Blc, and Tfu0937-Slp, the N terminus of each anchor protein was fused to the C terminus of Tfu0937. (C) E. coli BW25113; (D) E. coli JCM20137. All data are averages from three independent experiments, and error bars represent standard deviations.

Fig. 2.

Growth analysis of Tfu0937-displaying E. coli using cellobiose (A and B) or cellooligosaccharide (C and D) as the sole carbon source. (A) E. coli BW25113; (B) E. coli JCM20137. Symbols shown in panels A and B are as follows: Blc-Tfu0937 (closed circles), Tfu0937-Blc (open circles), and Tfu0937-Slp (triangles). Symbols shown in panels C and D are as follows: cellotriose (closed circles), cellotetraose (squares), cellopentaose (open circles), and cellohexaose (triangles). Data are averages from three independent experiments, and standard deviations were within 10%.