Insights into the action of phylogenetically diverse microbial expansins on the structure of cellulose microfibrils

Background Microbial expansins (EXLXs) are non-lytic proteins homologous to plant expansins involved in plant cell wall formation. Due to their non-lytic cell wall loosening properties and potential to disaggregate cellulosic structures, there is considerable interest in exploring the ability of microbial expansins (EXLX) to assist the processing of cellulosic biomass for broader biotechnological applications. Herein, EXLXs with different modular structure and from diverse phylogenetic origin were compared in terms of ability to bind cellulosic, xylosic, and chitinous substrates, to structurally modify cellulosic fibrils, and to boost enzymatic deconstruction of hardwood pulp. Results Five heterogeneously produced EXLXs ( Clavibacter michiganensis; Cmi EXLX2, Dickeya aquatica; Daq EXLX1, Xanthomonas sacchari; Xsa EXLX1, Nothophytophthora sp.; Nsp EXLX1 and Phytophthora cactorum; Pca EXLX1) were shown to bind xylan and hardwood pulp at pH 5.5 and Cmi EXLX2 (harboring a family-2 carbohydrate-binding module) also bound well to crystalline cellulose. Small-angle X-ray scattering revealed a 20–25% increase in interfibrillar distance between neighboring cellulose microfibrils following treatment with Cmi EXLX2, Daq EXLX1, or Nsp EXLX1. Correspondingly, combining xylanase with Cmi EXLX2 and Daq EXLX1 increased product yield from hardwood pulp by ~ 25%, while supplementing the Tr AA9A LPMO from Trichoderma reesei with Cmi EXLX2, Daq EXLX1, and Nsp EXLX1 increased total product yield by over 35%. Conclusion This direct comparison of diverse EXLXs revealed consistent impacts on interfibrillar spacing of cellulose microfibers and performance of carbohydrate-active enzymes predicted to act on fiber surfaces. These findings uncover new possibilities to employ EXLXs in the creation of value-added materials from cellulosic biomass.