Engineering <i>Leifsonia</i> Alcohol Dehydrogenase for Thermostability and Catalytic Efficiency by Enhancing Subunit Interactions

Zhu, Lu; Song, Yang; Chang, Chenchen; Ma, Hongmin; Yang, Lu; Deng, Zixin; Deng, Wei; Qu, Xudong

doi:10.1002/cbic.202100431

Cited by 8 publications

(7 citation statements)

References 24 publications

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“…In case of the tetrameric ADH from Leifsonia (LnADH) engineering of amino acids involved in the subunit contact areas proofed to be successful (Zhu et al, 2021).…”

Section: Protein Engineeringmentioning

confidence: 99%

Alcohol Dehydrogenases as Catalysts in Organic Synthesis

2022

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Alcohol dehydrogenases (ADHs) have become important catalysts for stereoselective oxidation and reduction reactions of alcohols, aldehydes and ketones. The aim of this contribution is to provide the reader with a timely update on the state-of-the-art of ADH-catalysis. Mechanistic basics are presented together with practical information about the use of ADHs. Current concepts of ADH engineering and ADH reactions are critically discussed. Finally, this contribution highlights some prominent examples and future-pointing concepts.

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“…In case of the tetrameric ADH from Leifsonia (LnADH) engineering of amino acids involved in the subunit contact areas proofed to be successful (Zhu et al, 2021).…”

Section: Protein Engineeringmentioning

confidence: 99%

Alcohol Dehydrogenases as Catalysts in Organic Synthesis

2022

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“…The development of in sillco design of enzyme reduce the experimental costs and time [9].These interfacial interactions are generally regulated by the key residues on the surface of motif of enzyme. Zhu et al [10] studied the modification of subunit interface of Leifsonia alcohol dehydrogenase (LnADH) and yielded the mutant T100R-S148I with thermostability with and catalytic efficiency enhancement. Boucher et al [11] investigated the structural basis for the hyperthermostability of a FN 3 -like protein domain from Ther-moanaerobacter tengcongensis by molecular dynamics simulations.…”

Section: Introductionmentioning

confidence: 99%

Design of Motifs Interfacial Interactions by Co-evolved Analysis of D-amino Acid Dehydrogenase for Stability Enhancement

Zhang

Duan

et al. 2023

Preprint

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To design D-amino acid dehydrogenase (DAADH) for enhanced stability, the interactions of the subunit interfaces of DAADH were analyzed. Interaction network analysis of DAADH indicated that there are only weak interactions between the A and B subunits. Several co-evolved residue pairs were selected for mutation to enhance interfacial interactions of subunits, and 11 designed MDHs were obtained. DA06 and DA11 were selected for experimental verification for their salt bridges are 1.4 and 1.2-fold of that of DAwild, respectively. DA11 can maintain 93% activity in 80℃, while it was only 40% for DAwild. Thermostabiliy study indicated the half-life of DA11 was 2-fold of DAwild. Molecular dynamics simulations revealed that the extraordinary stability of the DA11 was due to the formation of extra interfacial salt bridges. The paper provided a strategy of mutations outside the active site of enzyme by co-evolutionary analysis which can reduce the effect of the activity-thermostability trade-off.

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“…Generally, monooxygenases and oxidases have been developed for biocatalytic oxidation processes, and oxygen has been supplied as an electron acceptor (oxidant) with the ensuing H 2 O or H 2 O 2 -associated byproducts . Additionally, peroxygenases and dioxygenases have also been developed (although to a lesser extent), which makes the toolbox of the biocatalytic oxidation process even more diverse, but enzyme engineering is still required for enzyme activity and stability improvements and developing a broad substrate scope. , However, dehydrogenases are still attractive and relatively well developed with enhanced activity and thermostability . They are routinely used for reduction but can also be applied in biocatalytic oxidation processes together with the nicotinamide cofactor NAD(P) + as an electron acceptor.…”

Section: Introductionmentioning

confidence: 99%

“… 9 , 10 However, dehydrogenases are still attractive and relatively well developed with enhanced activity and thermostability. 11 They are routinely used for reduction but can also be applied in biocatalytic oxidation processes together with the nicotinamide cofactor NAD(P) + as an electron acceptor. Likewise, ( R )-undecavertol (>98%, ee ) was produced by ( S )-selective ADH alcohol dehydrogenase with a 400 g L –1 substrate concentration on a pilot scale.…”

Section: Introductionmentioning

confidence: 99%

Effect of Nitrogen, Air, and Oxygen on the Kinetic Stability of NAD(P)H Oxidase Exposed to a Gas–Liquid Interface

Wang

Erdem

Woodley

2023

Org. Process Res. Dev.

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Biocatalytic oxidation is an interesting prospect for the selective synthesis of active pharmaceutical intermediates. Bubbling air or oxygen is considered as an efficient method to increase the gas−liquid interface and thereby enhance oxygen transfer. However, the enzyme is deactivated in this process and needs to be further studied and understood to accelerate the implementation of oxidative biocatalysis in larger production processes. This paper reports data on the stability of NAD(P)H oxidase (NOX) when exposed to different gas−liquid interfaces introduced by N 2 (0% oxygen), air (21% oxygen), and O 2 (100% oxygen) in a bubble column. A pH increase was observed during gas bubbling, with the highest increase occurring under air bubbling from 6.28 to 7.40 after 60 h at a gas flow rate of 0.15 L min −1 . The kinetic stability of NOX was studied under N 2 , air, and O 2 bubbling by measuring the residual activity, the deactivation constants (k d1 ) were 0.2972, 0.0244, and 0.0346 with the corresponding half-lives of 2.2, 28.6, and 20.2 h, respectively. A decrease in protein concentration of the NOX solution was also observed and was attributed to likely enzyme aggregation at the gas−liquid interface. Most aggregation occurred at the air−water interface and decreased greatly from 100 to 14.16% after 60 h of bubbling air. Furthermore, the effect of the gas−liquid interface and the dissolved gas on the NOX deactivation process was also studied by bubbling N 2 and O 2 alternately. It was found that the N 2 −water interface and O 2 −water interface both had minor effects on the protein concentration decrease compared with the air−water interface, whilst the dissolved N 2 in water caused serious deactivation of NOX. This was attributed not only to the NOX unfolding and aggregation at the interface but also to the N 2 occupying the oxygen channel of the enzyme and the resultant inaccessibility of dissolved O 2 to the active site of NOX. These results shed light on the enzyme deactivation process and might further inspire bioreactor operation and enzyme engineering to improve biocatalyst performance.

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Engineering Leifsonia Alcohol Dehydrogenase for Thermostability and Catalytic Efficiency by Enhancing Subunit Interactions

Cited by 8 publications

References 24 publications

Alcohol Dehydrogenases as Catalysts in Organic Synthesis

Alcohol Dehydrogenases as Catalysts in Organic Synthesis

Design of Motifs Interfacial Interactions by Co-evolved Analysis of D-amino Acid Dehydrogenase for Stability Enhancement

Effect of Nitrogen, Air, and Oxygen on the Kinetic Stability of NAD(P)H Oxidase Exposed to a Gas–Liquid Interface

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