Influence of an accessory domain in biochemical and structural characteristics of the cellulase CelE2

Abstract

Different physicochemical and enzymatic treatments are applied in order to overcome the recalcitrance of plant cell wall to degradation. Glycosyl hydrolases (GHs) integrate a class of enzymes widely used in enzymatic process since they perform the hydrolysis of glycosidic bonds in a wide variety of polysaccharides. Apart from the catalytic domains, several glycosyl hydrolases contain accessories domains that can impact on biochemical and structural features. A previous study demonstrated that the cellulase CelE2, from metagenomic origin, contains a N-terminal domain characteristic of GH5 family and a second domain in the C-terminal region, displaying identity to Calx-beta domain, which is described as a cytoplasmic domain involved in exchange of ions sodium and calcium. Preliminary studies suggested the influence of C-terminal domain in CelE2 enzymatic activity. This domain had already been identified in other glycosyl hydrolases, however no clear function for this domain have been reported so far. In this context, the main objective of this project is to elucidate the influence of the C-terminal domain on CelE2 enzymatic activity. For this, biochemical and structural features will be evaluated through circular dichroism, small angle X-ray scattering experiments, as well as crystallographic and X-ray diffraction analysis. Furthermore, studies will be conduct to evaluate the ability of the C-terminal domain to bind to carbohydrates since the comparison with the dbCan database showed identity, although with low significance, with carbohydrate binding domains of family 35. Finally, biochemical and structural information will be used as input data for molecular dynamics studies aiming to access in details the probable molecular interactions occurring between the two domains of cellulase CelE2. Currently, there is no detailed information about the modular architecture of CelE2 and the elucidation of the hole of its C-terminal domain can provide support for the rational design of glycosyl hydrolases. (AU)