Engineering the activity of amine dehydrogenase in the asymmetric reductive amination of hydroxyl ketones

Ming, Hui; Yuan, Bo; Qu, Ge; Sun, Zhoutong

doi:10.1039/d2cy00391k

Cited by 8 publications

(5 citation statements)

References 66 publications

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“…Neither of the conversions showed any increas or decrease trends by elongating the reaction time. Using an ion exchange method, [24] the product NAS was obtained with a isolated yield of 80 % (8.73 g/L), giving 174.6 mg.…”

Section: Resultsmentioning

confidence: 99%

Engineering the Activity of a Newly Identified Arylalkylamine N‐Acetyltransferase in the Acetylation of 5‐Hydroxytryptamine

Wang,

Li,

Yuan

et al. 2024

ChemBioChem

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Arylalkylamine N‐acetyltransferase (AANAT) serves as a key enzyme in the biosynthesis of melatonin by transforming 5‐hydroxytryptamine (5‐HT) to N‐acetyl‐5‐hydroxytryptamine (NAS), while its low activity may hinder melatonin yield. In this study, a novel AANAT derived from Sus scrofa (SsAANAT) was identified through data mining using 5‐HT as a model substrate, and a rational design of SsAANAT was conducted in the quest to improving its activity. After four rounds of mutagenesis procedures, a triple combinatorial dominant mutant M3 was successfully obtained. Compared to the parent enzyme, the conversion of the whole‐cell reaction bearing the best variant M3 exhibted an increase from 50% to 99% in the transformation of 5‐HT into NAS. Additionally, its catalytic efficiency (kcat/Km) was enhanced by 2‐fold while retaining the thermostability (Tm > 45 °C). In the up‐scaled reaction with a substrate loading of 50 mM, the whole‐cell system incorporating variant M3 achieved a 99% conversion of 5‐HT in 30 h with an 80% yield. Molecular dynamics simulations were ultilized to shed light on the origin of improved activity. This study broadens the repertoire of AANAT for the efficient biosynthesis of melatonin.

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Section: Resultsmentioning

confidence: 99%

Engineering the Activity of a Newly Identified Arylalkylamine N‐Acetyltransferase in the Acetylation of 5‐Hydroxytryptamine

Wang,

Li,

Yuan

et al. 2024

ChemBioChem

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“…Activity towards hydroxyl-functionalized methyl ketones, studied by refs. 51 , 52 with engineered AmDHs, could be presumed based on the activity of Msme AmDH and Micro AmDH towards hexan-3-one in addition to 1-hydroxy-propan-2-one and 1-hydroxy-butan-2-one 53 .…”

Section: Resultsmentioning

confidence: 99%

A refined picture of the native amine dehydrogenase family revealed by extensive biodiversity screening

Elisée,

Ducrot,

Méheust

et al. 2024

Nat Commun

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Native amine dehydrogenases offer sustainable access to chiral amines, so the search for scaffolds capable of converting more diverse carbonyl compounds is required to reach the full potential of this alternative to conventional synthetic reductive aminations. Here we report a multidisciplinary strategy combining bioinformatics, chemoinformatics and biocatalysis to extensively screen billions of sequences in silico and to efficiently find native amine dehydrogenases features using computational approaches. In this way, we achieve a comprehensive overview of the initial native amine dehydrogenase family, extending it from 2,011 to 17,959 sequences, and identify native amine dehydrogenases with non-reported substrate spectra, including hindered carbonyls and ethyl ketones, and accepting methylamine and cyclopropylamine as amine donor. We also present preliminary model-based structural information to inform the design of potential (R)-selective amine dehydrogenases, as native amine dehydrogenases are mostly (S)-selective. This integrated strategy paves the way for expanding the resource of other enzyme families and in highlighting enzymes with original features.

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“…only 485 transformants, a 95% coverage was obtained. 61 Compared to conventional substrate-specificity engineering strategies used for amino acid dehydrogenases or amine dehydrogenases, such as rational site-directed mutagenesis 27,50,62,63 and iterative saturation mutagenesis (ISM), 29,31,64 our approach exhibits several significant benefits: first, the lower screening effort enables screening for more substrates in shorter times. Second, it allows the capture of potential synergistic effects arising from the combination of mutations at the five selected sites, an outcome that is nearly impossible to achieve by using traditional engineering strategies.…”

Section: Substrate-specific Evolution Of M0mentioning

confidence: 99%

“…Besides some chemical synthetic routes, − biocatalytic strategies have also been receiving considerable attention − mostly due to their high stereoselectivity and the mild reaction conditions. Particularly, amine dehydrogenase-catalyzed asymmetric reductive amination − and kinetic resolution − are two attractive processes to access chiral amines. Among them, the asymmetric reductive amination of α-hydroxy ketones by engineered amine dehydrogenase to yield β-amino alcohols has been extensively explored, achieving remarkable conversion rates and enantiomeric excess values of >99%. − Almost all known amine dehydrogenases exhibiting activity toward β-amino alcohols are mutants of natural l -amino acid dehydrogenases, exhibiting a strict ( S )-stereoselectivity.…”

Section: Introductionmentioning

confidence: 99%

Substrate-Specific Evolution of Amine Dehydrogenases for Accessing Structurally Diverse Enantiopure (R)-β-Amino Alcohols

Yin,

Gong,

Zeng

et al. 2024

ACS Catal.

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The biocatalytic oxidative deamination of β-amino alcohols holds significant practical potential in kinetic resolution and/or deracemization process to access (R)-β-amino alcohols. This study exemplifies a notable instance of acquisition and utilization of this valuable oxidative deamination activity. Initially, the mutation N261M (M0) was identified to endow a native valine dehydrogenase with oxidative deamination activity toward a few (S)-βamino alcohols. Subsequently, a phylogenetic analysis-guided, double-code saturation mutagenesis strategy was proposed to engineer M0's side-chain binding site. This strategy facilitated the substrate-specific evolution of M0, resulting in the creation of a panel of mutants (M1−M4) with noteworthy oxidative deamination activity toward structurally diverse (S)-β-amino alcohols. Using these engineered amine dehydrogenases, termed as β-amino alcohol dehydrogenases (β-AADHs), the complete kinetic resolution and even deracemization of a range of β-amino alcohols have been achieved. This work reports distinct biocatalysts and a synthetic strategy for the synthesis of enantiopure (R)-β-amino alcohols and offers an innovative approach for substrate-specificity engineering of enzymes.

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Engineering the activity of amine dehydrogenase in the asymmetric reductive amination of hydroxyl ketones

Abstract: Chiral amino alcholols are essential structural motifs required in pharmaceutical agents and bioactive natural products. Our previous work explored a series of engineered amine dehydrogenases (AmDHs) derived from amino acid...

Cited by 8 publications

References 66 publications

Engineering the Activity of a Newly Identified Arylalkylamine N‐Acetyltransferase in the Acetylation of 5‐Hydroxytryptamine

Engineering the Activity of a Newly Identified Arylalkylamine N‐Acetyltransferase in the Acetylation of 5‐Hydroxytryptamine

A refined picture of the native amine dehydrogenase family revealed by extensive biodiversity screening

Substrate-Specific Evolution of Amine Dehydrogenases for Accessing Structurally Diverse Enantiopure (R)-β-Amino Alcohols

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