A multienzyme biocatalytic cascade as a route towards the synthesis of α,ω-diamines from corresponding cycloalkanols

Sarak, Sharad; Pagar, Amol D.; Khobragade, Taresh P.; Jeon, Hyeongtag; Giri, Pritam; Lim, Seonga; Patil, Mahesh D.; Kim, Yechan; Kim, Byung Gee; Yun, Hyungdon

doi:10.1039/d2gc03392e

Cited by 6 publications

(5 citation statements)

References 78 publications

(27 reference statements)

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“…As mentioned above, pyruvate generated by glycolysis can be detrimental to transamination, leading to the deamination of the produced amine compounds. As a resolution of this, it was previously reported that point mutagenesis of an arginine residue in the active site of a transaminase causes loss of pyruvate acceptance, resulting in the improvement of short-chain amine compounds . To verify this, we first developed TA-PLP conjugate structures through homology modeling (Figure S14).…”

Section: Resultsmentioning

confidence: 99%

“…As a resolution of this, it was previously reported that point mutagenesis of an arginine residue in the active site of a transaminase causes loss of pyruvate acceptance, resulting in the improvement of short-chain amine compounds. 46 To verify this, we first developed TA-PLP conjugate structures through homology modeling (Figure S14). Next, we found target residues in interaction with the amine donor (benzylamine) and pyruvate, respectively, considering the possible hydrogen bonds.…”

Section: The Production Of αω-Diamines With Varying Carbon Chain Leng...mentioning

confidence: 99%

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One-Pot Biocatalytic Route from Alkanes to α,ω-Diamines by Whole-Cell Consortia of Engineered Yarrowia lipolytica and Escherichia coli

Kim,

Yoo,

Park

et al. 2024

ACS Synth. Biol.

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Metabolically engineered microbial consortia can contribute as a promising production platform for the supply of polyamide monomers. To date, the biosynthesis of long-chain α,ωdiamines from n-alkanes is challenging because of the inert nature of n-alkanes and the complexity of the overall synthesis pathway. We combined an engineered Yarrowia lipolytica module with Escherichia coli modules to obtain a mixed strain microbial consortium that could catalyze an efficient biotransformation of n-alkanes into corresponding α,ω-diamines. The engineered Y. lipolytica strain was constructed (YALI10) wherein the two genes responsible for β-oxidation and the five genes responsible for the overoxidation of fatty aldehydes were deleted. This newly constructed YALI10 strain expressing transaminase (TA) could produce 0.2 mM 1,12-dodecanediamine (40.1 mg/L) from 10 mM n-dodecane. The microbial consortia comprising engineered Y. lipolytica strains for the oxidation of n-alkanes (O M ) and an E. coli amination module (A M ) expressing an aldehyde reductase (AHR) and transaminase (TA) improved the production of 1,12-diamine up to 1.95 mM (391 mg/L) from 10 mM n-dodecane. Finally, combining the E. coli reduction module (R M ) expressing a carboxylic acid reductase (CAR) and an sfp phosphopantetheinyl transferase with O M and A M further improved the production of 1,12-diamine by catalyzing the reduction of undesired 1,12-diacids into 1,12-diols, which further undergo amination to give 1,12-diamine as the target product. This newly constructed mixed strain consortium comprising three modules in one pot gave 4.1 mM (41%; 816 mg/ L) 1,12-diaminododecane from 10 mM n-dodecane. The whole-cell consortia reported herein present an elegant "greener" alternative for the biosynthesis of various α,ω-diamines (C8, C10, C12, and C14) from corresponding n-alkanes.

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Section: Resultsmentioning

confidence: 99%

Section: The Production Of αω-Diamines With Varying Carbon Chain Leng...mentioning

confidence: 99%

One-Pot Biocatalytic Route from Alkanes to α,ω-Diamines by Whole-Cell Consortia of Engineered Yarrowia lipolytica and Escherichia coli

Kim,

Yoo,

Park

et al. 2024

ACS Synth. Biol.

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“…Notably, the only variable was the addition of glucose, which plays an important role in regenerating co-factors needed by CAR through the action of intracellular enzymes (Napora-Wijata et al, 2014). Since it is understood that glucose metabolism in resting whole cell biocatalysts generates substantial amounts of alternative amine acceptors such as pyruvate and ketoglutarate (Supporting Information: Figure S3a ) (Sarak et al, 2022), we looked at the influence of glucose concentration on the ability of WC2 to convert 1b. We saw that this was impaired at glucose concentrations of 55 mM and 110 mM (Supporting Information: Figure S3b ).…”

Section: Resultsmentioning

confidence: 99%

Reductive amination cascades in cell-free and resting whole cell formats for valorization of lignin deconstruction products

Nain,

Dickey,

Somasundaram

et al. 2023

Preprint

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The selective introduction of amine groups within deconstruction products of lignin could provide an avenue for valorizing waste biomass while achieving a green synthesis of industrially relevant building blocks from sustainable sources. Here, we built and characterized enzyme cascades that create aldehydes and subsequently primary amines from diverse lignin-derived carboxylic acids using a carboxylic acid reductase (CAR) and an ω-transaminase (TA). We report a detailed characterization of two reaction formats - cell-free biocatalysis and resting whole-cell biocatalysis - using the conversion of vanillate to vanillyl amine as model chemistry. We showed that resting whole-cell reactions could be competitive with cell-free reactions (97% and 70% yields, respectively) while avoiding steps of cell lysis, purification, and cofactor supplementation. We also demonstrated the recyclability of resting whole cells for multiple batches of catalysis without significant loss in productivity. Finally, we used the knowledge gained from this study to produce several amines from carboxylic acid precursors using one-pot biocatalytic reactions. These results expand our knowledge of these industrially relevant enzyme families to new substrates and contexts for environmentally friendly and potentially low-cost synthesis of diverse aryl aldehydes and amines.

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“…Notably, the only variable was the addition of glucose, which plays an important role in regenerating co‐factors needed by CAR through the action of intracellular enzymes (Napora‐Wijata et al, 2014). Since it is understood that glucose metabolism in resting whole cell biocatalysts generates substantial amounts of alternative amine acceptors such as pyruvate and ketoglutarate (Supporting Information S1: Figure 3a) (Sarak et al, 2022), we looked at the influence of glucose concentration on the ability of WC2 to convert 1b. We saw that this was impaired at glucose concentrations of 55 and 110 mM (Supporting Information S1: Figure 3d).…”

Section: Resultsmentioning

confidence: 99%

Reductive amination cascades in cell‐free and resting whole cell formats for valorization of lignin deconstruction products

Nain,

Dickey,

Somasundaram

et al. 2023

Biotech & Bioengineering

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The selective introduction of amine groups within deconstruction products of lignin could provide an avenue for valorizing waste biomass while achieving a green synthesis of industrially relevant building blocks from sustainable sources. Here, we built and characterized enzyme cascades that create aldehydes and subsequently primary amines from diverse lignin‐derived carboxylic acids using a carboxylic acid reductase (CAR) and an ω‐transaminase (TA). Unlike previous studies that have paired CAR and TA enzymes, here we examine multiple homologs of each of these enzymes and a broader set of candidate substrates. In addition, we compare the performance of these systems in cell‐free and resting whole‐cell biocatalysis formats using the conversion of vanillate to vanillyl amine as model chemistry. We also demonstrate that resting whole cells can be recycled for multiple batch reactions. We used the knowledge gained from this study to produce several amines from carboxylic acid precursors using one‐pot biocatalytic reactions, several of which we report for the first time. These results expand our knowledge of these industrially relevant enzyme families to new substrates and contexts for environmentally friendly and potentially low‐cost synthesis of diverse aryl aldehydes and amines.

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A multienzyme biocatalytic cascade as a route towards the synthesis of α,ω-diamines from corresponding cycloalkanols

Abstract: A multienzymatic cascade for the synthesis of 1,6-Hexamethylenediamine (HMD) and related α, ω-diamines from corresponding cycloalkanols is reported herein. In this work, an E coli-based modularization strategy was applied to...

Cited by 6 publications

References 78 publications

One-Pot Biocatalytic Route from Alkanes to α,ω-Diamines by Whole-Cell Consortia of Engineered Yarrowia lipolytica and Escherichia coli

One-Pot Biocatalytic Route from Alkanes to α,ω-Diamines by Whole-Cell Consortia of Engineered Yarrowia lipolytica and Escherichia coli

Reductive amination cascades in cell-free and resting whole cell formats for valorization of lignin deconstruction products

Reductive amination cascades in cell‐free and resting whole cell formats for valorization of lignin deconstruction products

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