Isobaric enzymatic assays reveal depth-dependent variation in microbial ectoenzyme activities in the deepest ocean on Earth

Microbial ectoenzymes play essential roles in the transformation and mineralization of organic matter in the ocean. However, conventional methods for measuring microbial ectoenzyme activities (MEAs) in the deep ocean under atmospheric conditions can misrepresent the metabolic activities of indigenous microbial communities. To overcome this limitation, we measured the activities of three microbial ectoenzymes (aminopeptidase, α-glucosidase, and β-glucosidase) under both atmospheric and in situ pressure conditions in seawater collected using pressure-retaining and non-pressure-retaining samplers in the Challenger Deep, Mariana Trench. In the case of aminopeptidase and α-glucosidase, the highest isobaric MEAs were observed in the surface layer (50 m), followed by those at abyssopelagic depths (4,000–6,000 m) for aminopeptidase, hadal realm (10,903 m) for α-glucosidase, while β-glucosidase activity exhibited the highest value at 10,903 m. Furthermore, the isobaric MEAs in hadal waters were commonly found to be higher than the decompressed counterparts, highlighting the importance of pressure-retaining sampling and isobaric enzymatic assays. The half-saturation constant (K_m) showed a general decreasing trend with depth, suggesting that the deep ocean microbes might have adapted to the high-pressure and oligotrophic environment by increasing their ectoenzyme’s affinity to substrate. Furthermore, particle-free MEAs contribute more to the total MEAs in the deep ocean than particle-associated MEAs, suggesting the significance of cell-associated and dissolved ectoenzymes in ocean ecosystems. This study provides a foundation for future investigations of MEAs in the ocean and has important implications for understanding the dynamics of microbially mediated biogeochemical cycling in marine ecosystems.