Dissecting and optimizing bioactivities of chitosans by enzymatic modification

Abstract

Chitosans are versatile biopolymers with antimicrobial and plant-strengthening properties relevant to agriculture. However, a limited understanding of molecular structure-function relationships and cellular modes of action of chitosans hampers the development of effective chitosan-based agro-biologics. We partially hydrolyzed a chitosan polymer (degree of polymerization DP 800, fraction of acetylation F_A 0.2) using acetic acid, a GH18 chitinase, or a GH8 chitosanase. All hydrolysates contained mixtures of chitosan oligomers and small polymers, but their composition in terms of DP, F_A, and pattern of acetylation (PA) differed greatly. Importantly, chitinase products had mostly deacetylated residues at their ends, flanking mostly deacetylated residues, and vice versa for chitosanase products, while the products of acid hydrolysis had random PA. Acid hydrolysis did not significantly change antifungal and antibacterial activities. In contrast, chitinase hydrolysis slightly increased antibacterial, and chitosanase almost abolished antifungal activity. Elicitor and priming activities in the plant Arabidopsis were unchanged by acid, destroyed by chitinase, and increased by chitosanase hydrolysis. Transcriptomic analysis revealed that the chitosan polymer strongly induced genes involved in photosynthesis, while the chitosanase hydrolysate strongly induced genes involved in disease resistance. Clearly, different bioactivities require different chitosans, and enzymatic modification can fine-tune these activities as required for different agricultural products.