The Reductive Amination of Carbonyl Compounds Using Native Amine Dehydrogenase from Laribacter hongkongensis

Lee, Somin; Jeon, Hyeongtag; Giri, Pritam; Lee, Uk‐Jae; Jung, Hyunsang; Lim, Seonga; Sarak, Sharad; Khobragade, Taresh P.; Kim, Byung‐Gee; Yun, Hyungdon

doi:10.1007/s12257-021-0113-2

Cited by 6 publications

(5 citation statements)

References 34 publications

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“…The variant wh81 was further used as template for the third round of mutagenesis. In this scenario, an important single mutation E114V was considered because it has been reported that this residue can function on the ammonia activation, and thereby affecting the enzyme activity ( Abrahamson et al, 2012 ; Chen et al, 2018 ; Patil et al, 2018 ; Lee et al, 2021 ). To our delight, the resultant quintuple variant wh84 (K68S/N261L/I111F/V294C/E114V) showed 99% conversion and >99% ee toward 1a.…”

Section: Resultsmentioning

confidence: 99%

Biosynthesis of Chiral Amino Alcohols via an Engineered Amine Dehydrogenase in E. coli

Tong

Qin

Wang

et al. 2022

Front. Bioeng. Biotechnol.

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Chiral amino alcohols are prevalent synthons in pharmaceuticals and synthetic bioactive compounds. The efficient synthesis of chiral amino alcohols using ammonia as the sole amino donor under mild conditions is highly desired and challenging in organic chemistry and biotechnology. Our previous work explored a panel of engineered amine dehydrogenases (AmDHs) derived from amino acid dehydrogenase (AADH), enabling the one-step synthesis of chiral amino alcohols via the asymmetric reductive amination of α-hydroxy ketones. Although the AmDH-directed asymmetric reduction is in a high stereoselective manner, the activity is yet fully excavated. Herein, an engineered AmDH derived from a leucine dehydrogenase from Sporosarcina psychrophila (SpAmDH) was recruited as the starting enzyme, and the combinatorial active-site saturation test/iterative saturation mutagenesis (CAST/ISM) strategy was applied to improve the activity. After three rounds of mutagenesis in an iterative fashion, the best variant wh84 was obtained and proved to be effective in the asymmetric reductive amination of 1-hydroxy-2-butanone with 4-fold improvements in kcat/Km and total turnover number (TTN) values compared to those of the starting enzyme, while maintaining high enantioselectivity (ee >99%) and thermostability (T5015 >53°C). In preparative-scale reaction, the conversion of 100 and 200 mM 1-hydroxy-2-butanone catalyzed by wh84 was up to 91–99%. Insights into the source of an enhanced activity were gained by the computational analysis. Our work expands the catalytic repertoire and toolbox of AmDHs.

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Section: Resultsmentioning

confidence: 99%

Biosynthesis of Chiral Amino Alcohols via an Engineered Amine Dehydrogenase in E. coli

Tong

Qin

Wang

et al. 2022

Front. Bioeng. Biotechnol.

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“…In 2021, Lee et al . discovered another new AmDH, LhAmDH, from Laribacter hongkongensis , which demonstrated excellent activity against cyclohexanone, achieving >99 % conversion with both whole cells and purified enzyme [70] . In 2022, Bennett et al .…”

Section: Biocatalysismentioning

confidence: 99%

“…In 2021, Lee et al discovered another new AmDH, LhAmDH, from Laribacter hongkongensis, which demonstrated excellent activity against cyclohexanone, achieving > 99 % conversion with both whole cells and purified enzyme. [70] In 2022, Bennett et al determined the structure of MATOUAmDH2, building on its earlier discovery, and unveiled the complex changes that AmDH undergoes during the catalytic cycle. [71] The enzymes mutants have biocatalytic potential in the synthesis of small chiral alkylamines and small amino alcohols.…”

Section: Amine Dehydrogenases (Amdhs)mentioning

confidence: 99%

Asymmetric Reductive Amination in Organocatalysis and Biocatalysis

Li¹,

Zhou²,

Meng³

et al. 2023

Eur J Org Chem

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This review summarizes the recent progress of organocatalytic and biocatalytic asymmetric reductive amination (ARA), a challenging but important topic for drug discovery and the pharmaceutical industry. At present, ARA can be divided into three categories: metal catalysis, organic catalysis, and biocatalysis. In the past decade, transition metal‐catalysed ARA has been well established. Organocatalytic ARA has emerged as a powerful alternative to metal‐catalysed ARA, the hydrogen sources used in organocatalytic ARA are usually Hantzsch esters, benzothiazolines, boranes, and hydrosilanes, which require Lewis base or phosphoric acid catalysts to activate them to give secondary chiral amines. It is worth mentioning that biocatalytic ARA has made remarkable progress in the last decade, amino acid dehydrogenases, amine dehydrogenases, opine dehydrogenases and imine reductases have been successfully used in ARA.

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“…In this study, we aimed to apply the substrate walking strategy to evolve AmDH for accessing a panel of bulky fusedring and linked-ring aryl ketones (Figure S2), which were difficult-to-aminate compounds by previously reported AmDH toolboxes. [29][30][31][34][35][36][37][38][39]62 However, given the diversity of the structure of target substrate panels, applying the conventional substrate walking stratey to evolve the individual acceptance of each target substrate implies large screening efforts, which are not desired in a user-friendly protein engineering workflow. 63−65 Toward this end, we proposed a modified strategy based on conventional substrate walking to quickly evolve the collective substrate acceptance of AmDH.…”

Section: ■ Introductionmentioning

confidence: 99%

“…In this study, we aimed to apply the substrate walking strategy to evolve AmDH for accessing a panel of bulky fused-ring and linked-ring aryl ketones (Figure S2), which were difficult-to-aminate compounds by previously reported AmDH toolboxes. − ,− , However, given the diversity of the structure of target substrate panels, applying the conventional substrate walking stratey to evolve the individual acceptance of each target substrate implies large screening efforts, which are not desired in a user-friendly protein engineering workflow. − Toward this end, we proposed a modified strategy based on conventional substrate walking to quickly evolve the collective substrate acceptance of AmDH. Focusing on the primary factor affecting the binding of the bulky substrates of AmDHs, steric hindrance, ,, we speculate that utilizing the positional migration of the benzene ring (the bulkiest structural motif in target substrate panels) to expand the substrate binding space step by step can meet the steric demands for the binding of the structurally diverse aryl ketone panels.…”

Section: Introductionmentioning

confidence: 99%

Shield Machine-like Substrate Walking Strategy-Based Pocket Engineering of F-Amine Dehydrogenase for Accessing Structurally Diverse Fused-Ring and Linked-Ring Aryl Ketones

Wu,

Xu,

Nie

et al. 2024

ACS Catal.

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Although amine dehydrogenases (AmDHs) are emerging as attractive biocatalysts for chiral amine synthesis, their synthetic application in structurally diverse arylamines remains challenging, given the limited substrate acceptance. Substrate walking is an effective coevolution strategy to confer targeted substrate acceptance to an enzyme through a stepwise mutagenesis landscape adaptation. Here, based on the conventional substrate walking strategy, we report a shield machine-like substrate walking strategy to quickly evolve F-BbAmDH from Bacillus badius for accessing the difficult-to-aminate fused-ring and linked-ring aryl ketones. A set of monoring aryl ketone homologues with the benzene ring located at the end of the side-chain and regularly extended carbon skeletons was rationally selected as the transition substrates. A superior mutant library with expanded target fused-ring and linked-ring aryl ketone acceptance was identified based on the activity and specificity enhancement of the transition substrates, enabling the synthesis of pharmaceuticals and bioactive compound-related arylamines with up to 94% yield and 99% ee (R) or 99:1 cis/trans. Structure-based computational results provided molecular insights into the source of the expanded substrate acceptance. Our work demonstrates a concise engineering workflow for the collective acceptance evolution of enzymes for structurally diverse substrate panels and has promising prospects in enzyme engineering.

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The Reductive Amination of Carbonyl Compounds Using Native Amine Dehydrogenase from Laribacter hongkongensis

Cited by 6 publications

References 34 publications

Biosynthesis of Chiral Amino Alcohols via an Engineered Amine Dehydrogenase in E. coli

Biosynthesis of Chiral Amino Alcohols via an Engineered Amine Dehydrogenase in E. coli

Asymmetric Reductive Amination in Organocatalysis and Biocatalysis

Shield Machine-like Substrate Walking Strategy-Based Pocket Engineering of F-Amine Dehydrogenase for Accessing Structurally Diverse Fused-Ring and Linked-Ring Aryl Ketones

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