Impact of scaffoldin mechanostability on cellulosomal activity

Lignocellulose is the most abundant renewable carbon source in the biosphere. However, the main bottleneck in its conversion to produce second generation biofuels is the saccharification step: the hydrolysis of lignocellulosic material into soluble fermentable sugars. Some anaerobic bacteria have developed an extracellular multi-enzyme complex called the cellulosome that efficiently degrades cellulosic substrates. Cellulosome complexes rely on enzyme-integrating scaffoldins that are large non-catalytic scaffolding proteins comprising several cohesin modules and additional functional modules that mediate the anchoring of the complex to the cell surface and the specific binding to its cellulosic substrate. It was proposed that mechanical forces may affect the cohesins positioned between the cell- and cellulose-anchoring points in the so-called connecting region. Consequently, the mechanical resistance of cohesins within the scaffoldin is of great importance, both to understand cellulosome function and as a parameter of industrial interest, to better mimic natural complexes through the use of the established designer cellulosome technology. Here we study how the mechanical stability of cohesins in a scaffoldin affects the enzymatic activity of a cellulosome. We found that when a cohesin of low mechanical stability is positioned in the connecting region of a scaffoldin, the activity of the resulting cellulosome is reduced as opposed to a cohesin of higher mechanical stability. This observation directly relates mechanical stability of the scaffoldin-borne cohesins to cellulosome activity and provides a rationale for the design of artificial cellulosomes for industrial applications, by incorporating mechanical stability as a new industrial parameter in the biotechnology toolbox.