Biomimetic strategy for constructing cellulosomal operons in .

BACKGROUND

Enzymatic conversion of lignocellulosic biomass into soluble sugars is a major bottleneck in the plant biomass utilization. Several anaerobic organisms cope these issues via multiple-enzyme complex system so called 'cellulosome'. Hence, we proposed a "biomimic operon" concept for making an artificial cellulosome which can be used as a promising tool for the expression of cellulosomal enzymes in .

RESULTS

According to the proteomic analysis of ATCC27405 induced by Avicel or cellobiose, we selected eight highly expressed cellulosomal genes including a scaffoldin protein gene (), a cell-surface anchor gene (), two exoglucanase genes ( and ), two endoglucanase genes ( and ), and two xylanase genes ( and ). Arranging these eight genes in two different orders, we constructed two different polycistronic operons using the ordered gene assembly in method. This is the first study to express the whole CipA along with cellulolytic enzymes in . Each operon was successfully expressed in RM125, and the protein complex assembly, cellulose-binding ability, thermostability, and cellulolytic activity were demonstrated. The operon with a higher xylanase activity showed greater saccharification on complex cellulosic substrates such as Napier grass than the other operon.

CONCLUSIONS

In this study, a strategy for constructing an efficient cellulosome system was developed and two different artificial cellulosomal operons were constructed. Both operons could efficiently express the cellulosomal enzymes and exhibited cellulose saccharification. This strategy can be applied to different industries with cellulose-containing materials, such as papermaking, biofuel, agricultural compost, mushroom cultivation, and waste processing industries.