83 research outputs found

    In Situ Acetaldehyde Synthesis for Carboligation Reactions

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    The enzyme 4-oxalocrotonate tautomerase (4-OT) can promis-cuously catalyze various carboligation reactions using acetalde-hyde as a nucleophile. However, the highly reactive nature ofacetaldehyde requires intricate handling, which can impede itsusage in practical synthesis. Therefore, we investigated threeenzymatic routes to synthesize acetaldehyde in situ in one-potcascade reactions with 4-OT. Two routes afforded practicalacetaldehyde concentrations, using an environmental pollu-tant,trans-3-chloroacrylic acid, or a bio-renewable, ethanol, asstarting substrate. These routes can be combined with 4-OTcatalyzed Michael-type additions and aldol condensations inone pot. This modular systems biocatalysis methodology pro-vides a stepping stone towards the development of larger arti-ficial metabolic networks for the practical synthesis of impor-tant chemical synthons

    Recent Applications of Carbon‐Nitrogen Lyases in Asymmetric Synthesis of Noncanonical Amino Acids and Heterocyclic Compounds

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    Carbon-nitrogen (C−N) lyases are enzymes that normally catalyze the cleavage of C−N bonds. Reversing this reaction towards carbon-nitrogen bond formation can be a powerful approach to prepare valuable compounds that could find applications in everyday life. This review focuses on recent (last five years) applications of native and engineered C−N lyases, either as stand-alone biocatalysts or as part of multienzymatic and chemoenzymatic cascades, in enantioselective synthesis of noncanonical amino acids and dinitrogen-fused heterocycles, which are useful tools for neurobiological research and important synthetic precursors to pharmaceuticals and food additives

    Enzymatic Oxy- and Amino-Functionalization in Biocatalytic Cascade Synthesis:Recent Advances and Future Perspectives

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    Biocatalytic cascades are a powerful tool for building complex molecules containing oxygen and nitrogen functionalities. Moreover, the combination of multiple enzymes in one pot offers the possibility to minimize downstream processing and waste production. In this review, we illustrate various recent efforts in the development of multi-step syntheses involving C-O and C-N bond-forming enzymes to produce high value-added compounds, such as pharmaceuticals and polymer precursors. Both in vitro and in vivo examples are discussed, revealing the respective advantages and drawbacks. The use of engineered enzymes to boost the cascades outcome is also addressed and current co-substrate and cofactor recycling strategies are presented, highlighting the importance of atom economy. Finally, tools to overcome current challenges for multi-enzymatic oxy- and amino-functionalization reactions are discussed, including flow systems with immobilized biocatalysts and cascades in confined nanomaterials.</p

    Biocatalytic Asymmetric Cyclopropanations via Enzyme‐bound Iminium Ion Intermediates

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    Cyclopropane rings are an important structural motif frequently found in many natural products and pharmaceuticals. Commonly, biocatalytic methodologies for the asymmetric synthesis of cyclopropanes rely on repurposed or artificial heme enzymes. Here, we engineered an unusual cofactor‐independent cyclopropanation enzyme based on a promiscuous tautomerase for the enantioselective synthesis of various cyclopropanes via the nucleophilic addition of diethyl 2‐chloromalonate to α,ÎČ‐unsaturated aldehydes. The engineered enzyme promotes formation of the two new carbon‐carbon bonds with excellent stereocontrol over both stereocenters, affording the desired cyclopropanes with high diastereo‐ and enantiopurity (d.r. up to 25:1; e.r. up to 99:1). Our results highlight the usefulness of promiscuous enzymes for expanding the biocatalytic repertoire for non‐natural reactions

    Tuning Enzyme Activity for Nonaqueous Solvents:Engineering an Enantioselective “Michaelase” for Catalysis in High Concentrations of Ethanol

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    Enzymes have evolved to function under aqueous conditions and may not exhibit features essential for biocatalytic application, such as the ability to function in high concentrations of an organic solvent. Consequently, protein engineering is often required to tune an enzyme for catalysis in non‐aqueous solvents. In this study, we have used a collection of nearly all single mutants of 4‐oxalocrotonate tautomerase, which promiscuously catalyzes synthetically useful Michael‐type additions of acetaldehyde to various nitroolefins, to investigate the effect of each mutation on the ability of this enzyme to retain its “Michaelase” activity in elevated concentrations of ethanol. Examination of this mutability landscape allowed the identification of two hotspot positions, Ser30 and Ala33, at which mutations are beneficial for catalysis in high ethanol concentrations. The “hotspot” position Ala33 was then randomized in a highly enantioselective, but ethanol‐sensitive 4‐OT variant (L8F/M45Y/F50A) to generate an improved enzyme variant (L8F/A33I/M45Y/F50A) that showed great ethanol stability and efficiently catalyzes the enantioselective addition of acetaldehyde to nitrostyrene in 40 % ethanol (permitting high substrate loading) to give the desired γ‐nitroaldehyde product in excellent isolated yield (89 %) and enantiopurity (ee=98 %). The presented work demonstrates the power of mutability‐landscape‐guided enzyme engineering for efficient biocatalysis in non‐aqueous solvents

    Macrophage migration inhibitory factor family proteins are multitasking cytokines in tissue injury

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    The family of macrophage migration inhibitory factor (MIF) proteins in humans consist of MIF, its functional homolog D-dopachrome tautomerase (D-DT, also known as MIF-2) and the relatively unknown protein named DDT-like (DDTL). MIF is a pleiotropic cytokine with multiple properties in tissue homeostasis and pathology. MIF was initially found to associate with inflammatory responses and therefore established a reputation as a pro-inflammatory cytokine. However, increasing evidence demonstrates that MIF influences many different intra- and extracellular molecular processes important for the maintenance of cellular homeostasis, such as promotion of cellular survival, antioxidant signaling, and wound repair. In contrast, studies on D-DT are scarce and on DDTL almost nonexistent and their functions remain to be further investigated as it is yet unclear how similar they are compared to MIF. Importantly, the many and sometimes opposing functions of MIF suggest that targeting MIF therapeutically should be considered carefully, taking into account timing and severity of tissue injury. In this review, we focus on the latest discoveries regarding the role of MIF family members in tissue injury, inflammation and repair, and highlight the possibilities of interventions with therapeutics targeting or mimicking MIF family proteins