Assembling mini-xylanosomes with Clostridium thermocellum XynA, and their properties in lignocellulose deconstruction.

Lignocellulose is a prominent source of carbohydrates to be used in biorefineries. One of the main challenges associated with its use is the low yields obtained during enzymatic hydrolysis, as well as the high cost associate with enzyme acquisition. Despite the great attention in using the fraction composed by hexoses, nowadays, there is a growing interest in enzymatic blends to deconstruct the pentose-rich fraction. Among the organisms studied as a source of enzymes to lignocellulose deconstruction, the anaerobic bacterium Clostridium thermocellum stands out. Most of the remarkable performance of C. thermocellum in degrading cellulose is related to its capacity to assemble enzymes into well-organized enzymatic complexes, cellulosomes. A mini-version of a cellulosome was designed in the present study, using the xylanase XynA and the N-terminus portion of scaffolding protein, mCipA, harboring one CBM3 and two cohesin I domains. The formed mini-xylanosome displayed maximum activity between 60 and 70 °C in a pH range from 6 to 8. Although biochemical properties of complexed/non-complexed enzymes were similar, the formed xylanosome displayed higher hydrolysis at 60 and 70 °C for alkali-treated sugarcane bagasse. Lignocellulose deconstruction using fungal secretome and the mini-xylanosome resulted in higher d-glucose yield, and the addition of the mCipA scaffolding protein enhanced cellulose deconstruction when coupled with fungal enzymes. Results obtained in this study demonstrated that the assembling of xylanases into mini-xylanosomes could improve sugarcane deconstruction, and the mCipA protein can work as a cellulose degradation enhancer.