Pinotsis, NikosNikosPinotsisKrüger, AnnaAnnaKrügerTomas, NicolasNicolasTomasChatziefthymiou, Spyros D.Spyros D.ChatziefthymiouLitz, ClaudiaClaudiaLitzMortensen, Simon ArnoldSimon ArnoldMortensenDaffé, MamadouMamadouDafféMarrakchi, HediaHediaMarrakchiAntranikian, GarabedGarabedAntranikianWilmanns, MatthiasMatthiasWilmanns2023-12-132023-12-132023-11-27Nature Communications 14 (1): 7649 (2023-11-27)https://hdl.handle.net/11420/44585The identification and characterization of enzyme function is largely lacking behind the rapidly increasing availability of large numbers of sequences and associated high-resolution structures. This is often hampered by lack of knowledge on in vivo relevant substrates. Here, we present a case study of a high-resolution structure of an unusual orphan lipase in complex with an endogenous C18 monoacylglycerol ester reaction intermediate from the expression host, which is insoluble under aqueous conditions and thus not accessible for studies in solution. The data allowed its functional characterization as a prototypic long-chain monoacylglycerol lipase, which uses a minimal lid domain to position the substrate through a hydrophobic tunnel directly to the enzyme’s active site. Knowledge about the molecular details of the substrate binding site allowed us to modulate the enzymatic activity by adjusting protein/substrate interactions, demonstrating the potential of our findings for future biotechnology applications.en2041-172320231Springerhttps://creativecommons.org/licenses/by/4.0/Chemical EngineeringJournal Article10.15480/882.892810.1038/s41467-023-43354-410.15480/882.8928Journal Article