Effect of multimodularity and spatial organization of glycoside hydrolases on catalysis.

The wide diversity among the carbohydrate-active enzymes (CAZymes) reflects the equally broad versatility in terms of composition and chemicals bonds found in the plant cell wall polymers on which they are active. This diversity is also expressed through the various strategies developed to circumvent the recalcitrance of these substrates to biological degradation. Glycoside hydrolases (GHs) are the most abundant of the CAZymes and are expressed as isolated catalytic modules or in association with carbohydrate-binding module (CBM), acting in synergism within complex arrays of enzymes. This multimodularity can be even more complex. The cellulosome presents a scaffold protein immobilized to the outer membrane of some microorganisms on which enzymes are grafted to prevent their dispersion and increase catalytic synergism. In polysaccharide utilization loci (PUL), GHs are also distributed across the membranes of some bacteria to co-ordinate the deconstruction of polysaccharides and the internalization of metabolizable carbohydrates. Although the study and characterization of these enzymatic activities need to take into account the entirety of this complex organization-in particular because of the dynamics involved in it-technical problems limit the present study to isolated enzymes. However, these enzymatic complexes also have a spatiotemporal organization, whose still neglected aspect must be considered. In the present review, the different levels of multimodularity that can occur in GHs will be reviewed, from its simplest forms to the most complex. In addition, attempts to characterize or study the effect on catalytic activity of the spatial organization within GHs will be addressed.

Potential of Ophiostoma piceae sterol esterase for biotechnologically relevant hydrolysis reactions.

The ascomycete Ophiostoma piceae produces a sterol esterase (OPE) with high affinity toward p-nitrophenol, glycerol, and sterol esters. Recently, this enzyme has been heterologously expressed in the methylotrophic yeast Pichia pastoris under the AOX1 methanol-inducible promoter (PAOX1) using sorbitol as co-susbtrate, and the hydrolytic activity of the recombinant protein (OPE*) turned out to be improved from a kinetic point of view. In this study, we analyze the effects of sorbitol during the expression of OPE*, at first added as an additional carbon source, and methanol as inducer. The O. piceae enzyme was successfully used for PVAc hydrolysis, suggesting its potential applicability in recycled paper production to decrease stickies problems.