Exploring the fungistatic mechanism of glycoside hydrolase 17 family endo-β-1,3-glucanase from Vitis vinifera for effectively inhibiting phytopathogens

A V. vinifera GH 17 β-1,3-glucanase was heterologously expressed and characterized. • Its catalytic groove accommodates β-1,6 branches, enabling antifungal action. • VvGlu17a produces β-1,3-oligosaccharides (DP 2–6) with prebiotic potential. • VvGlu17a exhibits potential in agricultural biocontrol and biomedical prebiotic production. β-1,3-Glucanases, a class of pathogenesis-related (PR) proteins, play critical roles in plant defense against phytopathogens. However, the enzymatic mechanism underlying their fungistatic effect remains to be further explored. This study focused on Vitis vinifera GH17 family endo -β-1,3-glucanase (VvGlu17a) to clarify its enzymatic properties and fungistatic mechanism. A genome-wide identification identified 54 GH17 β-1,3-glucanase genes in V. vinifera , with VvGlu17a belonging to subfamily B and possessing only a conserved GH17 domain. The purified recombinant VvGlu17a with optimal activity at pH 6.0 and 55 °C, and a specific activity of 408 U/mg toward linear β-1,3-glucan (curdlan), producing oligosaccharides with degrees of polymerization ranging from 2 to 6. In vitro fungistatic assays demonstrated that VvGlu17a significantly inhibited the growth of four phytopathogens, with the highest inhibition against Colletotrichum gloeosporioides . Structural modeling revealed that VvGlu17a adopts a typical (β/α)₈ TIM-barrel fold with an elongated, twisted catalytic groove containing eight glycosyl-binding sites (−3 to +5). Notably, the unobstructed C6 positions at subsites −1 to +3 allowing the enzyme to accommodate β-1,6-branched glucans—the structural feature of fungal cell wall polysaccharides. This study provides insights into the fungistatic mechanism of GH17 β-1,3-glucanases and their potential for developing biocontrol agents and functional oligosaccharides. Download: Download high-res image (259KB) Download: Download full-size image