Solubilization of crystalline chitin with a single LPMO: generating chito-oligosaccharides with unprecedented bioactivity

Abstract

Chitin is the second most abundant biopolymer in nature after cellulose and is very recalcitrant. The most valuable chitin-derived products are soluble polymeric chitosan and oligomeric chito-oligosaccharides, the production of which involves environment-unfriendly processing steps with concentrated acid and alkali. Enzymatic valorization of chitin has not been explored industrially, and enzymatic routes from chitin to soluble oligomeric or polymeric products are lacking. Lytic polysaccharide monooxygenases (LPMOs) contribute to chitin turnover in nature because they promote the activity of chitinases. Here, we have explored whether such LPMOs can be used as a stand-alone enzyme to solubilize chitin, yielding, in a single enzymatic step, bioactive oxidized chito-oligosaccharides (oxCHOS). We show that, indeed, chitin can be solubilized using only an LPMO, reaching up to 34 % solubilization with a single enzyme treatment and 60 % after four consecutive treatments. Importantly, the resulting soluble oxCHOS, with a degree of polymerization of 4 – 10, showed much stronger immunostimulatory activity on murine macrophages compared to non-oxidized CHOS. We also show that further diversification of the resulting oxCHOS may be achieved by subsequent treatment with chitin deacetylases. These results are poised to reshape current perceptions regarding the valorization of chitin biomass, opening a new green processing route, directly from chitin to products with high bioactivity.