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Title (Primary) Characterization of different biocatalyst formats for BVMO‐catalyzed cyclohexanone oxidation
Author Bretschneider, L.; Heuschkel, I.; Ahmed, A.; Bühler, K.; Karande, R.; Bühler, B.
Journal Biotechnology and Bioengineering
Year 2021
Department SOMA
Volume 118
Issue 7
Page From 2719
Page To 2733
Language englisch
Topic T7 Bioeconomy
Supplements https://onlinelibrary.wiley.com/action/downloadSupplement?doi=10.1002%2Fbit.27791&file=bit27791-sup-0001-Supplementary_Data.docx
Keywords Baeyer‐Villiger monooxygenase; biocatalysis; biofilm kinetics; enzyme kinetics; whole‐cell kinetics
Abstract Cyclohexanone monooxygenase (CHMO), a member of the Baeyer‐Villiger monooxygenase family, is a versatile biocatalyst that efficiently catalyzes the conversion of cyclic ketones to lactones. In this study, an Acidovorax‐derived CHMO gene was expressed in Pseudomonas taiwanensis VLB120. Upon purification, the enzyme was characterized in vitro and shown to feature a broad substrate spectrum and up to 100% conversion in 6 h. Further, we determined and compared the cyclohexanone conversion kinetics for different CHMO‐biocatalyst formats, i.e., isolated enzyme, suspended whole cells, and biofilms, the latter two based on recombinant CHMO‐containing P. taiwanensis VLB120. Biofilms showed less favorable values for KS (9.3‐fold higher) and kcat (4.8‐fold lower) compared to corresponding KM and kcat values of isolated CHMO, but a favorable KI for cyclohexanone (5.3‐fold higher). The unfavorable KS and kcat values are related to mass transfer‐ and possibly heterogeneity issues and deserve further investigation and engineering, in order to exploit the high potential of biofilms regarding process stability. Suspended cells showed an only 1.8‐fold higher KS, but 1.3‐ and 4.2‐fold higher kcat and KI values than isolated CHMO. This together with the efficient NADPH regeneration via glucose metabolism makes this format highly promising from a kinetics perspective.
Persistent UFZ Identifier https://www.ufz.de/index.php?en=20939&ufzPublicationIdentifier=24504
Bretschneider, L., Heuschkel, I., Ahmed, A., Bühler, K., Karande, R., Bühler, B. (2021):
Characterization of different biocatalyst formats for BVMO‐catalyzed cyclohexanone oxidation
Biotechnol. Bioeng. 118 (7), 2719 - 2733