Figure 1

Design of a minicellulosome structure on the yeast surface. (A) Schematic diagram of the overall concept of this research. The basic design of this research is composed of four different yeast strains, one for displaying the mini CipA, and the others for secreting three types of cellulases. Mini CipA, a modified scaffoldin including a CBM and three cohesin domains, was expressed as an Aga2-fusion protein. The dockerin fused-enzymes, C. thermocellum CelA and T. reesei CBHII, were randomly assembled to mini CipA via cohesin-dockerin interactions, whereas the secreted A. aculeatus BGLI was independently bound to the cell surface through its own cell surface adhesion characteristic. Coh, cohesin; CBM, carbohydrate-binding module; DocA and DocS, native dockerin in CelA and exogenous dockerin from C. thermocellum CelS, respectively. (B) Construction of a surface display vector, pCT-mini CipA. Mini CipA, a modified scaffoldin containing a CBM and three cohesin domains, was expressed under the control of GAL1 promoter as an Aga2-fusion protein for displaying on the yeast surface. (C) Construction of cellulase secretion vectors. α-factor prepro-peptide (s.s) was used as a signal sequence.