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Titel (primär) Characterization of different biocatalyst formats for BVMO‐catalyzed cyclohexanone oxidation
Autor Bretschneider, L.; Heuschkel, I.; Ahmed, A.; Bühler, K.; Karande, R.; Bühler, B.
Journal / Serie Biotechnology and Bioengineering
Erscheinungsjahr 2021
Department SOMA
Band/Volume 118
Heft 7
Seite von 2719
Seite bis 2733
Sprache englisch
Topic T7 Bioeconomy
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.
dauerhafte UFZ-Verlinkung
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