Phylogenetically distinct fungal expansins show different binding preferences towards cellulosic materials and enhance cellulase activity

Expansins and expansin-like proteins are found in plants and microbes, and can disrupt the cellulosic microfibril network of plant cell walls. While plant expansins play a role in cell wall formation, microbial expansin-like proteins reportedly enhance the activity of lignocellulolytic enzymes. Herein, two novel fungal expansin-like proteins, AmaEXLX1 from Allomyces macrogynus and ApuEXLX1 from Aureobasidium pullulans, were recombinantly produced in Pichia pastoris. While both AmaEXLX1 and ApuEXLX1 retain typical expansin structure, they share low sequence identity (22.5 %) and different predicted pI values (5.8 and 8.8, respectively), which was expected to impact their action on cellulosic substrates. Accordingly, adsorption of the proteins on cellulose nanofibrils (CNF) and the impact of the expansin-like proteins on the hydrolysis of CNF by an endoglucanase (Cel7B) was investigated using quartz crystal microbalance with dissipation (QCM-D). AmaEXLX1 showed higher affinity towards cellulose compared to ApuEXLX1, which was correlated to missing key aromatic residues in the polysaccharide binding surface of ApuEXLX1. The pretreatment of a CNF film with AmaEXLX1 and ApuEXLX1 increased the initial rate of Cel7B activity. This study underscores similarities between the impacts that bacterial and fungal expansin-like proteins can have on the enzymatic deconstruction of cellulose, and sequence properties that could impact expansin performance.