Kemp Elimination Catalyzed by Naturally Occurring Aldoxime Dehydratases

Miao, Yun‐gen; Metzner, Richard; Asano, Yasuhisa

doi:10.1002/cbic.201600596

Cited by 33 publications

(47 citation statements)

References 38 publications

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“…Kemp elimination (proton elimination from 5-nitrobenzisoxazole; Fig. 1 ) has been extensively used as a model system in enzyme design owing to the simplicity of the base-catalyzed ring opening reaction 11 and the absence of natural Kemp eliminases 1 , although some enzymes have been shown to catalyze Kemp elimination promiscuously 12 , 13 . Computational design of KE07 involved construction of a theozyme to catalyze the chemical reaction, which was then grafted into the scaffold of imidazole glycerol phosphate synthase (HisF) from Thermotoga maritima 1 .…”

Section: Introductionmentioning

confidence: 99%

The evolution of multiple active site configurations in a designed enzyme

et al. 2018

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Developments in computational chemistry, bioinformatics, and laboratory evolution have facilitated the de novo design and catalytic optimization of enzymes. Besides creating useful catalysts, the generation and iterative improvement of designed enzymes can provide valuable insight into the interplay between the many phenomena that have been suggested to contribute to catalysis. In this work, we follow changes in conformational sampling, electrostatic preorganization, and quantum tunneling along the evolutionary trajectory of a designed Kemp eliminase. We observe that in the Kemp Eliminase KE07, instability of the designed active site leads to the emergence of two additional active site configurations. Evolutionary conformational selection then gradually stabilizes the most efficient configuration, leading to an improved enzyme. This work exemplifies the link between conformational plasticity and evolvability and demonstrates that residues remote from the active sites of enzymes play crucial roles in controlling and shaping the active site for efficient catalysis.

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Section: Introductionmentioning

confidence: 99%

The evolution of multiple active site configurations in a designed enzyme

et al. 2018

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“…Although the Kemp elimination reaction is not catalyzed by any natural enzyme as a primary reaction, it can be catalyzed by several natural enzymes in a promiscuous way . In the first case, the ketoesteroid isomerases use the same mechanism as the already mentioned artificial enzymes .…”

Section: Introductionmentioning

confidence: 99%

“…[18] Although the Kemp elimination reaction is not catalyzed by any natural enzyme as a primary reaction, it can be catalyzed by several natural enzymes in a promiscuous way. [20][21][22][23] In the first case, the ketoesteroid isomerases use the same mechanism as the already mentioned artificial enzymes. [21] In the other two cases, the cytochrome P450 monooxygenase (P450-BM3) [22] and the Aldoxime dehydratases [23] use a redox mechanism.…”

Section: Introductionmentioning

confidence: 99%

“…[20][21][22][23] In the first case, the ketoesteroid isomerases use the same mechanism as the already mentioned artificial enzymes. [21] In the other two cases, the cytochrome P450 monooxygenase (P450-BM3) [22] and the Aldoxime dehydratases [23] use a redox mechanism. The electric fields in the active center of the enzymes and in solution can now be experimentally measured through vibrational Stark effect spectroscopy of specific groups.…”

Section: Introductionmentioning

confidence: 99%

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Kemp Elimination Reaction Catalyzed by Electric Fields

et al. 2020

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The Kemp elimination reaction is the most widely used in the de novo design of new enzymes. The effect of two different kinds of electric fields in the reactions of acetate as a base with benzisoxazole and 5‐nitrobenzisoxazole as substrates have been theoretically studied. The effect of the solvent reaction field has been calculated using the SMD continuum model for several solvents; we have shown that solvents inhibit both reactions, the decrease of the reaction rate being larger as far as the dielectric constant is increased. The diminution of the reaction rate is especially remarkable between aprotic organic solvents and protic solvents as water, the electrostatic term of the hydrogen bonds being the main factor for the large inhibitory effect of water. The presence of an external electric field oriented in the direction of the charge transfer (z axis) increases it and, so, the reaction rate. In the reaction of the nitro compound, if the electric field is oriented in an orthogonal direction (x axis) the charge transfer to the NO2 group is favored and there is a subsequent increase of the reaction rate. However, this increase is smaller than the one produced by the field in the z axis. It is worthwhile mentioning that one of the main effects of external electric fields of intermediate intensity is the reorientation of the reactants. Finally, the implications of our results in the de novo design of enzymes are discussed.

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“…Thus, opposite enantiomers are accessible starting from the same racemic aldehyde and the same enzyme. Taking such a broad substrate scope into account, for the future, a broad range of synthetic applications in organic nitrile synthesis can be expected with this novel biocatalytic approach, similar to the recently discovered activity for the Kemp elimination …”

Section: Discussionmentioning

confidence: 99%

Biocatalytic Synthesis of Nitriles through Dehydration of Aldoximes: The Substrate Scope of Aldoxime Dehydratases

et al. 2018

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Nitriles, which are mostly needed and produced by the chemical industry, play a major role in various industry segments, ranging from high-volume, low-price sectors, such as polymers, to low-volume, high-price sectors, such as chiral pharma drugs. A common industrial technology for nitrile production is ammoxidation as a gas-phase reaction at high temperature. Further popular approaches are substitution or addition reactions with hydrogen cyanide or derivatives thereof. A major drawback, however, is the very high toxicity of cyanide. Recently, as a synthetic alternative, a novel enzymatic approach towards nitriles has been developed with aldoxime dehydratases, which are capable of converting an aldoxime in one step through dehydration into nitriles. Because the aldoxime substrates are easily accessible, this route is of high interest for synthetic purposes. However, whenever a novel method is developed for organic synthesis, it raises the question of substrate scope as one of the key criteria for application as a "synthetic platform technology". Thus, the scope of this review is to give an overview of the current state of the substrate scope of this enzymatic method for synthesizing nitriles with aldoxime dehydratases. As a recently emerging enzyme class, a range of substrates has already been studied so far, comprising nonchiral and chiral aldoximes. This enzyme class of aldoxime dehydratases shows a broad substrate tolerance and accepts aliphatic and aromatic aldoximes, as well as arylaliphatic aldoximes. Furthermore, aldoximes with a stereogenic center are also recognized and high enantioselectivities are found for 2-arylpropylaldoximes, in particular. It is further noteworthy that the enantiopreference depends on the E and Z isomers. Thus, opposite enantiomers are accessible from the same racemic aldehyde and the same enzyme.

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Kemp Elimination Catalyzed by Naturally Occurring Aldoxime Dehydratases

Cited by 33 publications

References 38 publications

The evolution of multiple active site configurations in a designed enzyme

The evolution of multiple active site configurations in a designed enzyme

Kemp Elimination Reaction Catalyzed by Electric Fields

Biocatalytic Synthesis of Nitriles through Dehydration of Aldoximes: The Substrate Scope of Aldoxime Dehydratases

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