Marine microorganisms have evolved unique enzymatic repertoires under extreme conditions (high pressure, high salinity, and low temperature) offering promising biocatalysts for sustainable industrial biotechnology. However, translating these marine genetic resources into deployable solutions requires overcoming bottlenecks in enzyme discovery, mechanistic understanding, engineering, and process integration. Here, we present a systematic framework spanning from marine metagenome mining to pilot-scale bioprocess validation, focusing on three cascade biocatalytic systems addressing distinct waste-to-chemical conversion challenges.
     First, we established a sequence- and function-based high-throughput screening platform from over 20 marine metagenomic libraries (deep-sea, hydrothermal vents, cold seeps, and mangrove sediments), successfully retrieving six key enzymes: lipases, fatty acid photodecarboxylases, vanadium-dependent haloperoxidases, nitrilases, chitinases, and N-acetylglucosaminidases. Second, by integrating structural biology, molecular dynamics simulations, and AI-assisted design (AlphaFold3, Rosetta), we systematically elucidated the structure–function relationships of these marine enzymes. Guided by these mechanistic insights, we engineered enzyme variants with significantly enhanced catalytic efficiency and tailored substrate selectivity through rational and semi-rational design strategies.
      Based on these engineered enzymes, we constructed three cascade biocatalytic systems with distinct application potentials: (1) Lipase and photodecarboxylases cascade for converting algal oil-derived triglycerides into fatty acids and subsequently into alkanes, serving as next-generation biodiesel; (2) Haloperoxidases and nitrilase cascade for valorizing proteinaceous waste hydrolysates by converting L-glutamate into 3-cyanopropionic acid and further into succinic acid, which serves as a key monomer for poly(butylene succinate), a leading biodegradable plastic; and (3) Chitinase and N-acetylglucosaminidase cascade for depolymerizing marine shell waste chitin into N-acetylglucosamine monomers, a high-value additive in cosmetics, functional foods, and aquaculture feed.
     To bridge the gap between laboratory innovation and industrial deployment, we developed a plug-and-play modular immobilization system (SpyCatcher/Tag, DogCatcher/Tag) enabling flexible enzyme spacing and stoichiometry, along with co-expression strategies for two-enzyme cascades.
     This work demonstrates a complete translational pipeline, from genetic resource discovery and mechanistic dissection to engineered biocatalysts and scalable bioprocesses, offering ready-to-deploy enzymatic solutions for carbon-neutral synthesis, marine waste valorization, and sustainable ocean management.