Aspartic acid residues in BBE-like enzymes from <i>Morus alba</i> promote a function shift from oxidative cyclization to dehydrogenation
Berberine bridge enzyme (BBE)-like enzymes catalyze various oxidative cyclization and dehydrogenation reactions in natural product biosynthesis, but the molecular mechanism underlying the selectivity remains unknown. Here, we elucidated the catalytic mechanism of BBE-like oxidases from <i>Morus alba</i> involved in the oxidative cyclization and dehydrogenation of moracin C. X-ray crystal structures of a functionally promiscuous flavin adenine dinucleotide (FAD)-bound oxidase, MaDS1, with and without an oxidative dehydrogenation product were determined at 2.03 Å and 2.21 Å resolution, respectively. Structure-guided mutagenesis and sequence analysis have identified a conserved aspartic acid that directs the reaction toward the oxidative dehydrogenation pathway. A combination of density functional theory (DFT) calculations and molecular dynamics (MD) simulations has revealed that aspartic acid acts as the catalytic base to deprotonate the carbon-cation intermediate to generate the dehydrogenated product, which otherwise undergoes a spontaneous 6π electrocyclization in the oxidative cyclization pathway to furnish the 2H-benzopyran product.
Authors
- Junping Fan (ORCID: https://orcid.org/0000-0002-2615-8098)
- Xiaoguang Lei (ORCID: https://orcid.org/0000-0002-0380-8035)
- Jun Gu
- Kendall N. Houk (ORCID: https://orcid.org/0000-0002-8387-5261)
- Lei Gao (ORCID: https://orcid.org/0000-0003-3033-5285)
- Nianxin Guo (ORCID: https://orcid.org/0009-0004-7973-7490)
- Qixuan Wang (ORCID: https://orcid.org/0000-0001-6612-3634)
- Haoran Dong
- Dongshan Wu (ORCID: https://orcid.org/0000-0001-5049-8116)
- Qingyang Zhou (ORCID: https://orcid.org/0000-0002-3205-8579)
- Fang Liu (ORCID: https://orcid.org/0000-0002-0046-8434)
- Jun Yang (ORCID: https://orcid.org/0000-0002-2785-6971)
- Qi Ding
Institutions
- Nanjing Agricultural University (CN)
- University of California, Los Angeles (US)
- Peking University (CN)
- King Center (US)
- Nanjing University of Science and Technology (CN)
- Wuhan University (CN)
- Beijing National Laboratory for Molecular Sciences (CN)
- Cancer Research Institute of the Slovak Academy of Sciences (SK)
- Center for Interdisciplinary Studies (IN)
- China Shenhua Energy (China) (CN)
- Center for Life Sciences (CN)
- University of California, Berkeley (US)
Publication Details
- Journal
- Proceedings of the National Academy of Sciences
- Published
- 2025-08-19
- DOI
- https://doi.org/10.1073/pnas.2504346122
- Citations
- 1
- Primary Topic
- Enzyme Catalysis and Immobilization
- Type
- article
- Field-Weighted Citation Impact
- 0.81
Funders
- National Natural Science Foundation of China
- National Key Research and Development Program of China
- Beijing National Laboratory for Molecular Sciences