Colorimetric High-Throughput Screening Method for Directed Evolution of Prazole Sulfide Monooxygenase.
Chembiochem
; 23(16): e202200228, 2022 08 17.
Article
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| MEDLINE
| ID: mdl-35639013
ABSTRACT
Baeyer-Villiger monooxygenases (BVMOs) are important biocatalysts for the enzymatic synthesis of chiral sulfoxides, including chiral sulfoxide-type proton pump inhibitors for the treatment of gastrointestinal diseases. However, native BVMOs are not yet suitable for practical application due to their unsatisfactory activity and thermostability. Although protein engineering approaches can help address these issues, few feasible high-throughput methods are available for the engineering of such enzymes. Herein, a colorimetric detection method to distinguish sulfoxides from sulfides and sulfones was developed for prazole sulfide monooxygenases. Directed evolution enabled by this method has identified a prazole sulfide monooxygenase CbBVMO variant with improved activity and thermostability that catalyzes the asymmetric oxidation of lansoprazole sulfide. A 71.3 % increase in conversion and 6 °C enhancement in the melting point were achieved compared with the wild-type enzyme. This new method is feasible for high-throughput screening of prazole sulfide monooxygenase variants with improved activity, thermostability, and/or substrate specificity.
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Texto completo:
1
Banco de datos:
MEDLINE
Asunto principal:
Ensayos Analíticos de Alto Rendimiento
/
Oxigenasas de Función Mixta
Tipo de estudio:
Diagnostic_studies
/
Screening_studies
Idioma:
En
Revista:
Chembiochem
Asunto de la revista:
BIOQUIMICA
Año:
2022
Tipo del documento:
Article