Synthesized Research Articles

Structural Insights and Reaction Profile of a New Unspecific Peroxygenase from Marasmius wettsteinii Produced in a Tandem-Yeast Expression System.

Fungal unspecific peroxygenases (UPOs) are gaining momentum in synthetic chemistry. Of special interest is the UPO from Marasmius rotula (MroUPO), which shows an exclusive repertoire of oxyfunctionalizations, including the terminal hydroxylation of alkanes, the α-oxidation of fatty acids and the C-C cleavage of corticosteroids. However, the lack of heterologous expression systems to perform directed evolution has impeded its engineering for practical applications. Here, we introduce a close ortholog of MroUPO, a UPO gene from Marasmius wettsteinii (MweUPO-1), that has a similar reaction profile to MroUPO and for which we have set up a directed evolution platform based on tandem-yeast expression. Recombinant MweUPO-1 was produced at high titers in the bioreactor (0.7 g/L) and characterized at the biochemical and atomic levels. The conjunction of soaking crystallographic experiments at a resolution up to 1.6 Å together with the analysis of reaction patterns sheds light on the substrate preferences of this promiscuous biocatalyst.

Hydrazone-Functionalized trans-A2B-Corroles: Effective Synergy in Photodynamic Therapy of Lung Cancer.

A synthetic route to trans-A2B-corroles combining the macrocyclic core with a hydrazone moiety, based on the reactivity of azoalkenes toward dipyrromethanes, has been established with the aim of developing a new class of photosensitizers for photodynamic therapy of lung cancer. The study of the photophysical properties of the novel macrocycles allowed the identification of photosensitizers with absorption within the phototherapeutic window and high singlet oxygen quantum yield. Relevant structure-photodynamic activity correlations were established by studying the new corroles-based photodynamic therapy (PDT) in human lung cancer cell lines (A549 and H1299). The methyl-hydrazone corroles were more active than phenyl-hydrazone corroles, with the N-Boc and N-Ts groups being key structural features to ensure high activity. The lead photosensitizers, with IC50 values below 100 nM and no cytotoxicity per se, were significantly more active than 5,10,15-triphenylcorrole, showing that the presence of the hydrazone functional group has a strong influence on PDT activity.

Chalcogen bonds enable efficient photoreduction of sulfur-containing heterocycles.

Chalcogen bonding interactions have attracted significant attention in a broad chemistry community, with a particular focus on their ability to stabilise the key transition states in various organic synthetic routes. In this work, we demonstrate that they can also be harnessed in selective photoredox reactions, which cannot be otherwise achieved with alternative approaches to photoreduction. We demonstrate this concept through the photoreduction of the sulfur-containing DNA nucleoside precursor thioanhydrouridine to 2'-deoxy-thiouridine, revealing the previously unrecognized role of bisulfide in this process. Based on quantum chemical simulations, we identify a stable chalcogen-bonding complex of the hydrosulfide anion and thionhydrouridine (HS-…S contacts), which enables directional photoinduced electron transfer, resulting in the formation of non-canonical DNA nucleoside. We also disprove the possibility that photoreduction of thioanhydronucleosides could be initiated by hydrated electrons generated from irradiated bisulfide anions which do not interact with the chromophore. Finally, we show that selective photoreduction mediated by chalcogen bonds can only occur for chromophores, which exhibit sufficiently long excited-state lifetimes in the locally-excited states to undergo transition to the productive charge transfer state. These findings can be further used in the design of similar photoredox reactions which can employ the potential of chalcogen bonding interactions.

Promotion of RNF168-Mediated Nucleosomal H2A Ubiquitylation by Structurally defined K63-Polyubiquitylated Linker Histone H1.

The chemical synthesis of histones with homogeneous modifications is a potent approach for quantitatively deciphering the functional crosstalk between different post-translational modifications (PTMs). Here, we developed an expedient site-specific (poly)ubiquitylation strategy (CAEPL, Cysteine-Aminoethylation coupled with Enzymatic Protein Ligation), which integrates the Cys-aminoethylation reaction with the process of ubiquitin-activating enzyme UBA1-assisted native chemical ligation. Using this strategy, we successfully prepared monoubiquitylated and K63-linked di- and tri-ubiquitylated linker histone H1.0 proteins, which were incorporated into individual chromatosomes. Quantitative biochemical analysis of different RNF168 constructs on ubiquitylated chromatosomes with different ubiquitin chain lengths demonstrated that K63-linked polyubiquitylated H1.0 could directly stimulate RNF168 ubiquitylation activity by enhancing the affinity between RNF168 and chromatosome. Subsequent cryo-EM structural analysis of the RNF168/UbcH5c-Ub/H1.0-K63-Ub3 chromatosome complex revealed the potential recruitment orientation between RNF168 UDM1 domain and K63-linked ubiquitin chain on H1.0. Finally, we explored the impact of H1.0 ubiquitylation on RNF168 activity in the context of asymmetric H1.0-K63-Ub3 di-nucleosome substrate, revealing a comparable stimulation effect of both the inter- and intra-nucleosomal crosstalk. Overall, our study highlights the significance of access to structurally-defined polyubiquitylated H1.0 by CAEPL strategy, enabling in-depth mechanistic investigations of in-trans PTM crosstalk between linker histone H1.0 and core histone H2A ubiquitylation.

Toward Sustainable Chemical Synthesis: Metathesis of Sunflower-Derived Ethyl Esters and Evaluation of the Surfactant Potential of the Product Mixtures

Toward Sustainable Chemical Synthesis: Metathesis of Sunflower-Derived Ethyl Esters and Evaluation of the Surfactant Potential of the Product Mixtures

Equimolar Cross-Coupling Using Reactive Coiled Coils for Covalent Protein Assemblies.

Biocompatible cross-coupling reactions enable the efficient covalent attachment of large biomolecules at near-stoichiometric ratios, ensuring the stability and integrity of the resulting products. We present an affinity-based peptide platform utilizing coiled coils containing reactive side chains for proximity-driven protein cross-coupling in the presence of a cross-linking agent. This platform supports both chemical synthesis and recombinant expression, using canonical amino acids to generate reactive affinity tags. Employing the E3/R3 coiled coil pair as a scaffold, we design four complementary coils with cysteine residues as cross-linking sites, achieving >90% conversion to covalent heterodimeric coupling products using 3,4-dibromomaleimide. Equimolar mixtures of proteins with reactive coils at their termini yield near-quantitative heterodimeric cross-coupling products. The strategic selection of complementary coiled coil pairs and cross-linking agents enables orthogonal assembly of macromolecules with diverse architectures. This method offers a versatile approach for creating covalent fusion proteins, enhancing their stability and functionality for applications in chemical biology, biotechnology, and medicine.

Effect of the ZrO2 Crystal Phase on Performance over a CuCeZrO Catalyst for Dehydrogenative Condensation of Ethanol to Ethyl Acetate.

Ethyl acetate is an important chemical, and ethanol dehydrogenative condensation with 95.6% atomic economy is a promising synthetic route. CuZrO-based catalysts are widely applied in this reaction; however, the effect of the ZrO2 crystal phase in the CuCeZrO catalyst upon reaction deserves to be explored further. Herein, the ZrO2 crystal phase in the CuCeZrO catalyst was regulated by controlling the calcination temperature, and a high temperature (≥600 °C) is conducive to the transformation of the tetragonal phase to the monoclinic phase. Additionally, monoclinic ZrO2 (m-ZrO2) with limited density of the strong base is found to be more beneficial to the selective production of ethyl acetate from ethanol, while tetragonal ZrO2 (t-ZrO2) with abundant strongly basic sites is more inclined to the formation of butanol and its downstream esters. This work not only improves our understanding of the crystal phase effect in a CuCeZrO catalytic system but also paves the way for the development of more efficient catalysts for ethyl acetate production.

Stereoselective Synthesis of Chiral C2-Symmetric 1,3- and 1,5-Bis-Sulfoxides Guided by the Horeau Principle: Understanding the Influence of the Carbon Chain Nature in Its Ability for Metal Coordination.

The stereoselective synthesis of two distinct types of C2-symmetric chiral bis-sulfoxides, 1,3- and 1,5-bis(sulfinyl) derivatives, has been achieved based on the DAG methodology. The 1,5-bis(sulfinyl) derivatives constitute a new family of tridentate chiral ligands thanks to the presence of an additional sulfenyl or sulfinyl group in the carbon chain acting as a bridge. A systematic development and optimization of two synthetic routes, one for each ligand family, have been undertaken, highlighting the strategic utilization of Horeau's law to enhance enantioselectivity. Additionally, palladium (Pd) and ruthenium (Ru) complexes derived from the synthesized bis-sulfoxides were prepared, and their structures were elucidated through spectroscopic analysis. Isolation of Pd(II) complexes involving 1,3-bis-sulfoxides was exclusively achieved using trifluoroacetates as coligands. In the case of Ru(II) complexes, the trans geometry could be determined for 1,3-bis-sulfoxides. The introduction of a third sulfur atom as a coordinating element in the 1,5-bis(sulfinyl) derivatives facilitates the formation of two distinct tricoordinated Ru(II) complexes. The structure of these complexes is intricately influenced by the oxidation state adopted by the central sulfur on the chain, whether as a thioether or as a sulfoxide.

Polyoxometalate-embedded copper-organic network as dual-site synergetic catalyst for the selective oxidation of benzylic C−H bond

Selective activation and oxidization of inert C−H bonds into value-added chemicals affords attractively economic and ecological benefits, as well as central challenge in modern synthetic chemistry. Herein, by integrating the redox polyoxometalate (POM) centers and multinuclear copper sites in one system, four compounds were constructed with formulas of [Cu2(μ2-Cl)(H2O)2L3][PM12O40]·4H2O (M = Mo for 1, W for 2), and [Cu4(μ2-OH)4L2] [H3PM11CuO40]·12H2O (M = Mo for 3, W for 4, L = 1,4-di[4H-1,2,4-triazol-4-yl]-benzene). Both compounds 1–2 exhibited 3-D POM-embedded supramolecular networks constructed by discrete [PM12O40]3− clusters and binuclear {Cu2(μ2-Cl)(N–N)2} unit-based metal-organic sheets. While compounds 3–4 displayed 3-D POM-supported supramolecular networks composed of monosubstituted Keggin-type anions [H3PM11CuO40]4− and {Cu2(μ2-OH)(N–N)2} chain-based metal-organic layers. Four crystalline structures with distinct dual-active site distribution showed significant peroxidase-like activities owing to the collaborative catalysis of multinuclear copper units and polyanionic clusters, which can smoothly catalyze the C−H bond oxidation of diphenylmethane to benzophenone with ca. 97 % conversion and >98 % selectivity within 24 h. Notably, the monosubstituted [H3PM11CuO40]4−-based compounds 3−4 showed higher catalytic activities than [PM12O40]3−-based compounds 1−2 (88 % conversion and >97 % selectivity within 12 h). This kind of crystalline structures also showed good recoverability. The synergetic reaction mechanism was also explored. This work provides a new strategy to design robust heterogeneous POM-based synergetic catalysts for C−H bond functionalization.

Possibilities of using secondary plant metabolites as antitumor agents

The review summarized the literature data of recent years on the antitumor effect of secondary plant metabolites, as well as their immunotropic and anti-inflammatory effects as components of the antitumor response. The biological basis for the action of secondary plant metabolites was characterized in the form of influence on potential targets: transcription factors, signaling pathways and receptors responsible for proliferation and apoptosis. The ways of increasing the bioavailability of secondary plant metabolites to enhance the effectiveness and possibility of their medicinal use were considered, the effects of berberine, curcumin and their derivatives were described. The search for scientific publications was conducted in foreign (PubMed) and domestic (eLibrary) electronic libraries. It was found that the multiplicity of molecular targets of secondary plant metabolites and the pleiotropy of their effects suggest the possibility of their use for the regulation of various processes in tumor and normal cells. There was a connection between the antitumor effect of secondary plant metabolites and their anti-inflammatory and immunomodulatory action. However, a significant limitation of their use was the fact that most studies were conducted on cell cultures, which was insufficient to judge the antitumor effect. Clinical trials were few and their results were contradictory. In addition, a certain contradiction has been noted between the idea of a more effective action when using a pure substance or a complex composition of various plant components. An important problem was the low bioavailability of most secondary plant metabolites, for which various methods have been proposed. Despite the long history of phytotherapy in oncology, the development of new derivatives of secondary plant metabolites with high water solubility remains relevant, including modified molecules of known secondary plant metabolites and the search for new ones with unexplored biological activity. Modern methods of chemical synthesis and delivery systems of derivatives of secondary plant metabolites, as well as the study of their effects in model experiments, seem to be promising scientific directions for the creation of new drugs with antitumor activity.

Total Synthesis of Ryanodane Diterpenoids Garajonone and 3‐epi‐Garajonone

Ryanodane diterpenes are structurally complex natural products that are well‐known for their high degree of oxidation and the challenges associated with synthesizing them within the terpene class. Herein, we present a two‐stage synthetic strategy that draws inspiration from the broad biosynthesis of terpenes, allowing us to successfully achieve the first chemical synthesis of garajonone, a ryanodane diterpenoid that occurs naturally at low abundance, as well as its epimer, 3‐epi‐garajonone. The key to this success lies in the rapid construction of the carbon framework of target molecule by employing an early‐stage palladium‐catalyzed Heck/carbonylative esterification cascade annulation, followed by successive late‐stage selective redox manipulation to establish the desired oxidation state of the molecule. This research not only showcases the synthesis of garajonone and its epimer but also provides a platform for the chemical synthesis of other members and analogs within this complex diterpenoid family.

Biomimetic folding strategies for chemical synthesis of disulfide-bonded peptides and proteins.

Disulfide-bonded peptides and proteins, including hormones, toxins, growth factors, and others, are abundant in living organisms. These molecules play crucial physiological roles such as regulating cell and organism growth, development, and metabolism. They have also found widespread applications as drugs or tool molecules in biomedical and pharmaceutical research. However, the chemical synthesis of disulfide-bonded proteins is complicated by the challenges associated with their folding. This review focuses on the latest advancements in disulfide-bonded peptide and protein folding technologies. Particularly, it highlights biomimetic folding strategies that emulate the naturally occurring oxidative folding processes in nature. These strategies include chaperone-assisted folding, glycosylation-assisted folding, and organic-based oxidative folding methods. The review also anticipates future directions in folding technology. Such research offers innovative approaches for the chemical synthesis of complex proteins that are otherwise difficult-to-fold.

Total Synthesis of Ryanodane Diterpenoids Garajonone and 3-epi-Garajonone.

Ryanodane diterpenes are structurally complex natural products that are well-known for their high degree of oxidation and the challenges associated with synthesizing them within the terpene class. Herein, we present a two-stage synthetic strategy that draws inspiration from the broad biosynthesis of terpenes, allowing us to successfully achieve the first chemical synthesis of garajonone, a ryanodane diterpenoid that occurs naturally at low abundance, as well as its epimer, 3-epi-garajonone. The key to this success lies in the rapid construction of the carbon framework of target molecule by employing an early-stage palladium-catalyzed Heck/carbonylative esterification cascade annulation, followed by successive late-stage selective redox manipulation to establish the desired oxidation state of the molecule. This research not only showcases the synthesis of garajonone and its epimer but also provides a platform for the chemical synthesis of other members and analogs within this complex diterpenoid family.

BracketMaker: Visualization and optimization of chemical protein synthesis.

Chemical protein synthesis (CPS), in which custom peptide segments of ~20-60 aa are produced by solid-phase peptide synthesis and then stitched together through sequential ligation reactions, is an increasingly popular technique. The workflow of CPS is often depicted with a "bracket" style diagram detailing the starting segments and the order of all ligation, desulfurization, and/or deprotection steps to obtain the product protein. Brackets are invaluable tools for comparing multiple possible synthetic approaches and serve as blueprints throughout a synthesis. Drawing CPS brackets by hand or in standard graphics software, however, is a painstaking and error-prone process. Furthermore, the CPS field lacks a standard bracket format, making side-by-side comparisons difficult. To address these problems, we developed BracketMaker, an open-source Python program with built-in graphic user interface (GUI) for the rapid creation and analysis of CPS brackets. BracketMaker contains a custom graphics engine which converts a text string (a protein sequence annotated with reaction steps, introduced herein as a standardized format for brackets) into a high-quality vector or PNG image. To aid with new syntheses, BracketMaker's "AutoBracket" tool automatically performs retrosynthetic analysis on a set of segments to draft and rank all possible ligation orders using standard native chemical ligation, protection, and desulfurization techniques. AutoBracket, in conjunction with an improved version of our previously reported Automated Ligator (Aligator) program, provides a pipeline to rapidly develop synthesis plans for a given protein sequence. We demonstrate the application of both programs to develop a blueprint for 65 proteins of the minimal Escherichia coli ribosome.

In Silico ADME And Molecular Docking Studies of New Thiazolyl-bipyrazole, Pyrazolopyridine and Pyrano[2,3-d]pyrazolopyridine Derivatives as Antibacterial Agents

: In this study, a series of novel pyrazole-based compounds were synthesized starting from the precursor ethyl 3-(4-amino-1-phenyl-3-((4-sulfamoylphenyl)carbamoyl)-1Hpyrazol- 5-yl)-3-oxopropanoate (2). Various synthetic routes were used to obtain pyrazolylpyrazolone 3, tricyclic dipyrazolopyridine 4a-c, thiazolyl-bipyrazoles 5 & 6, pyrazolo[4,3- b]pyridines 7 & 9, and tricyclic pyranopyrazolopyridine 10a–c. These compounds were screened for their antibacterial activity against four bacterial strains. The promising candidates 4a, 4b, 4c, 7, 9, and 10c exhibited minimum inhibitory concentrations ranging from 0.98 to 31.25 μg/mL. The in silico ADME properties for the active compounds exhibited similar physiochemical properties, with compound 9 demonstrating the best likeness and no inhibition effect on the popular drug metabolism enzyme CYP. Molecular docking simulations highlighted compounds 9 and 10c as potent antibacterial agents via DNA-gyrase inhibition

New biologically active sulfonamides as potential drugs for Alzheimer's disease.

A family of new compounds with sulfonamide and amide functional groups as potential Alzheimer's disease drugs were prepared by multistep synthesis. Thermal stability measurements recorded the initial decomposition in the range of 200-220°C, close above the melting point. The final compounds were tested for their ability to inhibit acetylcholinesterase and butyrylcholinesterase, and the in vitro dissolution behavior of selected compounds was studied through both lipophilic and hydrophilic matrix tablets. All nine tested derivatives were even more active in inhibiting acetylcholinesterase than the clinically used rivastigmine. Regression analysis of the obtained dissolution profiles was performed, and the effects of the pH and the release mechanism were determined. Some substances showed remarkable biological activity and became a subject of interest for further extensive study.

Indium‐Catalyzed Direct Amidation Reaction of Carboxylic Acids and In Silico Study for Screening the Activity of Potential Therapeutics of the Synthesized Products

AbstractAmide bonds are ubiquitous and valuable motifs in synthetic chemistry, found in a wide range of applications such as the backbone of proteins and pharmaceutical agents. Thus, environmentally friendly and selective methods for synthesizing amide bonds are important. Herein, we report a simple, efficient, and rapid route to access a range of functionalized amides from non‐activated carboxylic acids and amines using indium (III) trifluoromethanesulfonate as an efficient catalyst. A wide range of carboxylic acids – including aliphatic, aromatic, heterocyclic, and dicarbonyl acids – participate smoothly in these reactions, generating structurally diverse amides in moderate to good yields (up to 91 % in 24 examples). The reactions are conducted in dry tetrahydrofuran (THF) under reflux conditions. Furthermore, this amidation strategy provides a method for addressing challenging molecules, such as protected amino acids, to produce biocompatible products. These products can then be used as substrates for various organic transformations, including C−H functionalization reactions. We also demonstrate the synthetic utility of our protocol through the synthesis of essential substrates in organic and medicinal chemistry by reacting N‐protected carboxylic acids with 8‐aminoquinoline to produce a series of biologically valuable compounds. Five of these products, named 4 f, 4p, 9a, 9c, and 9d, have been biologically evaluated through an in‐silico study. The interaction of these compounds with receptor protein 8DQT was examined to assess their potential as drug candidates for hypertension treatment. Effective binding was detected between these compounds and the receptor. Data from molecular docking studies show that compounds 4d and 4 m are the most active, with binding scores of −8.8 and −8.6 kcal/mol, respectively. However, compound 4 m exhibited a stronger binding affinity than compound 4d over the course of dynamic simulation. Our ADMET study suggests that all five compounds are highly safe in the body. Based on the BOILED‐Egg model study, good absorption is suggested for all the molecules, and they can cross the blood‐brain barrier.

Protein‐Polymer Conjugates: Advancing Enzyme Catalysis in Synthetic Chemistry

Enzyme catalysis is prominent in chemical synthesis due to its specificity, selectivity, and efficiency. However, enzymes often face challenges, such as inactivation and inhibition in practical applications, and are also limited by narrow reaction scopes in complex catalytic scenarios like cascade reactions. These persistent issues have driven the development of chemical modifications of enzymes, aiming to enhance enzyme catalysis with external chemical entities. Polymers are particularly notable among these entities for their functional moieties and protective effects on biomolecules, enhancing enzyme properties and even creating new‐to‐nature catalysis by harnessing catalytic promiscuity. This concept aims to introduce the field of protein‐polymer conjugates for diverse emerging applications across emulsion biocatalysis, artificial enzymes, and supramolecular enzyme catalysis.

Molecular mechanics studies of factors affecting overall rate in cascade reactions: Multi-enzyme colocalization and environment.

Millions of years of evolution have optimized many biosynthetic pathways by use of multi-step catalysis. In addition, multi-step metabolic pathways are commonly found in and on membrane-bound organelles in eukaryotic biochemistry. The fundamental mechanisms that facilitate these reaction processes provide strategies to bioengineer metabolic pathways in synthetic chemistry. Using Brownian dynamics simulations, here we modeled intermediate substrate transportation of colocalized yeast-ester biosynthesis enzymes on the membrane. The substrate acetate ion traveled from the pocket of aldehyde dehydrogenase to its target enzyme acetyl-CoA synthetase, then the substrate acetyl CoA diffused from Acs1 to the active site of the next enzyme, alcohol-O-acetyltransferase. Arranging two enzymes with the smallest inter-enzyme distance of 60 Å had the fastest average substrate association time as compared with anchoring enzymes with larger inter-enzyme distances. When the off-target side reactions were turned on, most substrates were lost, which suggests that native localization is necessary for efficient final product synthesis. We also evaluated the effects of intermolecular interactions, local substrate concentrations, and membrane environment to bring mechanistic insights into the colocalization pathways. The computation work demonstrates that creating spatially organized multi-enzymes on membranes can be an effective strategy to increase final product synthesis in bioengineering systems.

Synthesis of New Indazole Analogs of Curcumin as Antioxidant and Anti-inflammatory Candidates: An <i>In Vitro</i> Investigation

The development of analog curcumin compounds by modifying the structure of monocarbonyl into an analog indazole of curcumin (AIC) is recognized to have a great potential. Still, only a few reports have been available. Rarely occurring in nature, indazole molecules are typically created through chemical synthesis. Therefore, this study aimed to synthesize six new AIC compounds with a particular focus on testing in vitro antioxidant activity using the DPPH and FRAP methods, as well as anti-inflammatory activity using the protein denaturation method. The results showed that the compounds formed had high anti-inflammatory activity but low antioxidant activity. All synthesis products produced higher anti-inflammatory activity than standard diclofenac sodium and curcumin compounds. Specifically, compound 3a showed the highest anti-inflammatory activity with an IC50 = 0.548 ± 0.062 μM. Therefore, it was concluded that compound 3a has the potential to be further studied for anti-inflammatory activity.