A novel carboxylesterase from Acinetobacter sp. JNU9335 for efficient biosynthesis of Edoxaban precursor with high substrate to catalyst ratio. (December 2020)

• Record Type: Journal Article
• Authors: Dou, Zhe; Xu, Guochao; Ni, Ye
• Is Part Of: Bioresource technology. Volume 317(2020)
• Publication Date: 2020-12
• Subjects: Carboxylesterase -- High-throughput screening -- Genome shotgun library, kinetic resolution -- Methyl 3-cyclohexene-1-carboxylate -- Substrate to catalyst ratio; Biomass -- Periodicals; Biomass energy -- Periodicals; Bioremediation -- Periodicals; Agricultural wastes -- Periodicals; Factory and trade waste -- Periodicals; Organic wastes -- Periodicals; Bioénergie -- Périodiques; Déchets agricoles -- Périodiques; Déchets industriels -- Périodiques; Déchets organiques -- Périodiques; Déchets (Combustible) -- Périodiques; 662.88
• DOI: 10.1016/j.biortech.2020.123984
• Languages: English
• ISSNs: 0960-8524

Abstract

Graphical abstract: Highlights: A sensitive and reliable HTS method was established to identify CHCM-hydrolyzing enzymes. Ac Est1 is a novel carboxylesterase with low sequence identity to known enzymes. Ac Est1 displays extremely high k cat and enantioselectivity toward CHCM. High S/C and substrate tolerance in kinetic resolution of CHCM. Abstract: A novel carboxylesterase Ac Est1 was identified from Acinetobacter sp. JNU9335 with high efficiency in the biosynthesis of chiral precursor of Edoxaban through kinetic resolution of methyl 3-cyclohexene-1-carboxylate (CHCM). Sequence analysis revealed Ac Est1 belongs to family IV of esterolytic enzymes and exhibits <40% identities with known carboxylesterases. The optimum pH and temperature of recombinant Ac Est1 are 8.0 and 40 °C. Substrate spectrum analysis indicated that Ac Est1 prefers substrates with short acyl and alcohol groups. Ac Est1 was highly active in the hydrolysis of CHCM with k cat of 1153 s −1 and displayed high substrate tolerance. As much as 2.0 M (280 g·L −1 ) CHCM could be enantioselectively hydrolyzed into ( S )-CHCM by merely 0.08 g·L −1 Ac Est1 with ee s of >99% ( S ) and substrate to catalyst ratio (S/C) of 3500 g·g −1 . These results indicate that the novel Ac Est1 is a promising biocatalyst in the synthesis of chiral carboxylic acids.