Expression and purification of recombinants xylanolitics enzymes and study of their activities against sugar cane extracted arabinoxylans

Abstract

The search for methods that use lignocellulosic biomass for production of second generation ethanol has lead to the study of enzymes able to hydrolyse this substrate and provide fermentable sugars. However, the heterogenic and recalcitrant composition of lignocellulosic biomass require a group of enzymes acting synergistically for its complete hydrolysis. This works aims to express, purify and characterize the activity of the enzymes Feruloyl Esterase (Fae), Acetyl Xylan Esterase (Axe), Arabinofuranosidase (Ara), Xylanase (Xyl) and alpha-Glucuronidase (AGlu), which have as substrate different glycosidic bonds on the polysaccharide arabinoxylan, part of that complex feedstock. pET-28 plasmids containing the enzymes insert will be transformed in Escherichia coli strains. The enzymes will be purified by affinity chromatography and used in the activity essays, alone and combined, against wheat and sugar cane arabinoxylan. Mass spectrometry will be used for detection of structures released after enzymatic treatment and imaging of the polymer surface, thus aiming to understand the architecture of the plant cell wall and the mechanism of synergistic action of lignocellulolytic enzymes and the possibility of elaborating a mix of enzymes that could work synergistically and efficiently hydrolyse lignocellulosic biomass. (AU)