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Du, Y.* ; Lv, X.* ; Siebenmorgen, T. ; Popowicz, G.M. ; Feng, C.* ; Ma, Y.* ; Wang, Y.*

Enhancing catalytic activity of a Baeyer-Villiger monooxygenase from Oceanicola granulosus: Simultaneous engineering of the distal site and active site.

J. Agric. Food Chem. 73, 14453-14466 (2025)
DOI PMC
Open Access Green as soon as Postprint is submitted to ZB.
Enzymatic synthesis of δ-lactones using Baeyer-Villiger monooxygenases (BVMOs) has potential in the fragrance and flavor industries but is constrained by poor activity toward ortho-alkyl-substituted cyclopentanones, key δ-lactone precursors. We determined the crystal structure of a BVMO derived from Oceanicola granulosus (OgBVMO), uncovering a unique loop adjacent to key catalytic residue R335. Site-saturation mutagenesis of loop residues A338 and A339 identified the A339E variant, obtaining a 2.4- to 3-fold increase in catalytic activity toward ortho-alkyl-substituted cyclopentanones (2-methyl-, 2-ethyl-, 2-hexyl-, and 2-heptylcyclopentanone). Further engineering the substrate-binding pocket yielded the Q442N variant, improving activity by 2.7-3.8-fold. Remarkably, the combinatorial mutant A339E/Q442N achieved 3.3-5.2-fold higher activity than the wild-type. Molecular dynamics simulations indicated that these improvements were driven by more favorable nucleophilic attack distances, underscoring the synergistic effects of distal and active-site mutations. These findings offer valuable insights into enhancing the catalytic activity of BVMOs, supporting the green manufacturing of high-value flavor compounds.
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Publication type Article: Journal article
Document type Scientific Article
Keywords Baeyer−villiger Monooxygenase ; Active Site ; Catalytic Activity ; Crystal Structure ; Distal Site ; Molecular Dynamics Simulation; Crystal-structure; Asymmetric Hydrogenation; Rational Design; Reduction; Lactones; Flavors
ISSN (print) / ISBN 0021-8561
e-ISSN 1520-5118
Journal Journal of Agricultural and Food Chemistry
Quellenangaben Volume: 73, Issue: 23, Pages: 14453-14466 Article Number: , Supplement: ,
Publisher American Chemical Society (ACS)
Publishing Place 1155 16th St, Nw, Washington, Dc 20036 Usa
Reviewing status Peer reviewed
Institute(s) Institute of Structural Biology (STB)
Grants State Key Laboratory of Pulp and Paper Engineering
Key Program of Natural Science Foundation of China
Key Realm R&D Program of GuangDong Province