A multi-dimensional engineering strategy integrating computational design, self-cyclization and whole cell biocatalysis for efficient epilactose production

Résumé fourni par la source

(CsCE) showed the highest epilactose synthesis activity. We therefore developed an integrated strategy combining computational design, SpyTag/SpyCatcher-mediated cyclization, and ethanol-permeabilized whole-cell catalysis. By combining enzyme ligand binding energy analysis, protein stability prediction, and catalytic constant prediction, the V52N variant was obtained. Its epilactose synthesis activity was 3.45 times that of the wild type, while lactulose formation was reduced to 14.8% of the wild-type level. Cyclized CCT increased the optimum temperature to 80 °C and extended the half-life at 85 °C by 5.52-fold. The optimized whole-cell process produced 65.81 g/L epilactose from 200 g/L lactose within 20 min, corresponding to 32.90% conversion. This strategy provides a practical route for efficient epilactose biomanufacturing.

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