Unnatural Derivatives Research Articles

Sustainable Chemical Derivatization of Hydroxycinnamic Acids

AbstractGreen solvents like ethyl acetate and acetone convert hydroxycinnamic acids into natural and unnatural derivatives such as amides, esters, and other valuable synthetic intermediates. Employing as protection‐activation a system based on an alkyl chloroformate and N‐methyl morpholine, ethyl acetate emerges as an excellent solvent for coupling O‐carbonate‐protected mixed anhydrides in acylation reactions of C‐protected amino acids, amines, and catalyzed acylation of alcohols. This results in high yields of O‐protected phenolic amides and esters. Acetone is preferred for acyl nucleophilic substitution reactions involving ionic species like free amino acids, sodium azide, and sodium hydroxide, all added in water. This approach releases the resultant O‐protected acylation products. Free phenolic compounds are promptly obtained on demand through carbonate deprotection. This synthetic method offers a simplified procedure and employs low‐cost reagents to prepare natural and unnatural hydroxycinnamic acid derivatives.

Substrate-Selective Catalysis Enabled Synthesis of Azaphilone Natural Products.

Achieving substrate-selectivity is a central element of nature's approach to synthesis. By relying on the ability of a catalyst to discriminate between components in a mixture, control can be exerted over which molecules will move forward in a synthesis. This approach can be powerful when realized but can be challenging to duplicate in the laboratory. In this work, substrate-selective catalysis is leveraged to discriminate between two intermediates that exist in equilibrium, subsequently directing the final cyclization to arrive at either the linear or angular tricyclic core common to subsets of azaphilone natural products. By using a flavin-dependent monooxygenase (FDMO) in sequence with an acyl transferase (AT), the conversion of several orcinaldehyde substrates directly to the corresponding linear tricyclic azaphilones in a single reaction vessel was achieved. Further, mechanistic studies support that a substrate equilibrium together with enzyme substrate selectivity play an import role in the selectivity of the final cyclization step. Using this strategy, five azaphilone natural products were synthesized for the first time as well as a number of unnatural derivatives thereof.

Controllable skeletal reorganizations in natural product synthesis.

Covering: 2016 to 2023The synthetic chemistry community is always in pursuit of efficient routes to natural products. Among the many available general strategies, skeletal reorganization, which involves the formation, cleavage, and migration of C-C and C-heteroatom bonds, stands out as a particularly useful approach for the efficient assembly of molecular skeletons. In addition, it allows for late-stage modification of natural products for quick access to other family members or unnatural derivatives. This review summarizes efficient syntheses of steroid, terpenoid, and alkaloid natural products that have been achieved by means of this strategy in the past eight years. Our goal is to illustrate the strategy's potency and reveal the spectacular human ingenuity demonstrated in its use and development.

Engineered Biosynthesis of Complex Disorazol Polyketides in a Streamlined Burkholderia thailandensis.

Engineering the biosynthetic pathways of complex natural products is a significant approach to obtain derivatives with improved properties. Here, we constructed a streamlined engineered biosynthesis system of myxobacterium-derived complex polyketide disorazol in a heterologous host, Burkholderia thailandensis E264. Inactivation of dehydratase domains in the disorazol biosynthetic pathway led to the production of two hydroxylated derivatives. Module deletion allowed the generation of an unnatural derivative with a truncated macrolactone ring, and the ACP-KS linker was the optimal fusion region for module deletion in this trans-AT polyketide synthase. These disorazol derivatives showed different activities against human cancer cell lines ranging from the nanomolar to micromolar level, suggesting the primary structure-activity relationship. The PKS engineering enables structural derivatization of disorazol, facilitating the in-depth engineered biosynthesis of polyketides.

Improved protein glycosylation enabled heterologous biosynthesis of monoterpenoid indole alkaloids and their unnatural derivatives in yeast

With over 3000 reported structures, monoterpenoid indole alkaloids (MIAs) constitute one of the largest alkaloid groups in nature, including the clinically important anticancer drug vinblastine and its semi-synthetic derivatives from Catharanthus roseus (Madagascar’s periwinkle). With the elucidation of the complete 28-step biosynthesis for anhydrovinblastine, it is possible to investigate the heterologous production of vinblastine and other medicinal MIAs. In this study, we successfully expressed the flavoenzyme O-acetylstemmadenine oxidase in Saccharomyces cerevisiae (baker’s yeast) by signal peptide modification, which is a vinblastine biosynthetic gene that has not been functionally expressed in this system. We also reported the simultaneous integration of ∼18 kb MIA biosynthetic gene cassettes as single copies into four genomic loci of baker’s yeast by CRISPR-Cas9, which enabled the biosynthesis of vinblastine precursors catharanthine and tabersonine from the feedstocks secologanin and tryptamine. We further demonstrated the biosynthesis of fluorinated and hydroxylated catharanthine and tabersonine derivatives using our yeasts, which showed that the MIA biosynthesis accommodates unnatural substrates, and the system can be further explored to produce other complex MIAs.

Role of a conserved ion-binding site tyrosine in ion selectivity of the Na+/K+ pump.

The essential transmembrane Na+ and K+ gradients in animal cells are established by the Na+/K+ pump, a P-type ATPase that exports three Na+ and imports two K+ per ATP hydrolyzed. The mechanism by which the Na+/K+ pump distinguishes between Na+ and K+ at the two membrane sides is poorly understood. Crystal structures identify two sites (sites I and II) that bind Na+ or K+ and a third (site III) specific for Na+. The side chain of a conserved tyrosine at site III of the catalytic α-subunit (Xenopus-α1 Y780) has been proposed to contribute to Na+ binding by cation-π interaction. We substituted Y780 with natural and unnatural amino acids, expressed the mutants in Xenopus oocytes and COS-1 cells, and used electrophysiology and biochemistry to evaluate their function. Substitutions disrupting H-bonds impaired Na+ interaction, while Y780Q strengthened it, likely by H-bond formation. Utilizing the non-sense suppression method previously used to incorporate unnatural derivatives in ion channels, we were able to analyze Na+/K+ pumps with fluorinated tyrosine or phenylalanine derivatives inserted at position 780 to diminish cation-π interaction strength. In line with the results of the analysis of mutants with natural amino acid substitutions, the results with the fluorinated derivatives indicate that Na+-π interaction with the phenol ring at position 780 contributes minimally, if at all, to the binding of Na+. All Y780 substitutions decreased K+ apparent affinity, highlighting that a state-dependent H-bond network is essential for the selectivity switch at sites I and II when the pump changes conformational state.

Application of Nucleic Acid Frameworks in the Construction of Nanostructures and Cascade Biocatalysts: Recent Progress and Perspective

Nucleic acids underlie the storage and retrieval of genetic information literally in all living organisms, and also provide us excellent materials for making artificial nanostructures and scaffolds for constructing multi-enzyme systems with outstanding performance in catalyzing various cascade reactions, due to their highly diverse and yet controllable structures, which are well determined by their sequences. The introduction of unnatural moieties into nucleic acids dramatically increased the diversity of sequences, structures, and properties of the nucleic acids, which undoubtedly expanded the toolbox for making nanomaterials and scaffolds of multi-enzyme systems. In this article, we first introduce the molecular structures and properties of nucleic acids and their unnatural derivatives. Then we summarized representative artificial nanomaterials made of nucleic acids, as well as their properties, functions, and application. We next review recent progress on constructing multi-enzyme systems with nucleic acid structures as scaffolds for cascade biocatalyst. Finally, we discuss the future direction of applying nucleic acid frameworks in the construction of nanomaterials and multi-enzyme molecular machines, with the potential contribution that unnatural nucleic acids may make to this field highlighted.

Structural and Mechanistic Insights into Chain Release of the Polyene PKS Thioesterase Domain

Polyketides serve as rich source of therapeutically relevant drug leads. The manipulation of polyketide synthases (PKSs) for generating derivatives with improved activities usually results in substantially reduced yields. Growing evidence suggests that type I PKS thioesterase (TE) domains are key bottlenecks in the biosynthesis of polyene antibiotics, such as pimaricin and amphotericin, and their unnatural derivatives. Herein, we elucidate the structure of the 26-membered macrolide-complexed TE domain from the pimaricin pathway (Pim TE), which specifies a spacious bifunnel-shaped substrate channel with a highly hydrophobic cleft proximal to the catalytic triad and a hydrophilic loop I region specific for the cyclization of amphiphilic polyene macrolide. Notably, the natural intermediate with C12-COOH is stabilized by a hydrogen-bond network, as well as by interactions between the polyene moiety and the hydrophobic cleft. Moreover, the bottleneck in processing the unnatural intermediate with C12-CH3 is attributed to the unstable and mismatched docking of the curved substrate in the channel. Aided by an in vitro assay with a fully elongated linear polyene intermediate as the substrate, multiple strategies were adopted, herein, to engineer Pim TE, including introducing H-bond donors, enhancing hydrophobic interactions, and modifying the catalytic center. Efficient TE mutations with increased substrate conversion up to 39.2% in vitro were further conducted in vivo, with a titer increase as high as 37.1% for the less toxic decarboxylated pimaricin derivatives with C12-CH3. Our work uncovers the mechanism of TE-catalyzed polyene macrolide formation and highlights TE domains as targets for PKS manipulation for titer increases in engineered unnatural polyketide derivatives.

Synthesis and biological evaluation of 10-benzyloxy-Narciclasine

A chemoenzymatic convergent synthesis of 10-benzyloxy narciclasine from bromobenzene was accomplished in 16 steps. The key transformations included toluene dioxygenase-mediated hydroxylation, nitroso Diels-Alder reaction and intramolecular Heck cyclization. The unnatural derivative of narciclasine was subjected to biological evaluation and its activity was compared to other C-10 and C-7 compounds prepared previously.

From Glycals to Nitrogen Heterocycles and Carbocycles via "Cleavage-Intramolecular Recombination Strategy".

Glycals (carbohydrate enol-ethers) have enjoyed profound applications in organic synthesis for more than a century. They not only serve as versatile glycosyl donors or as substrates for Ferrier rearrangement, but also find extensive synthetic applications especially as a "chiral pool" for accomplishing the synthesis of a variety of natural and biologically important compounds. As cyclic enol ethers, they demonstrate high reactivity and are among the most and variously transformable monosaccharide derivatives. The uniqueness of the reactivity of glycals is that they can be synthetically tuned to get a library of derivatives through stereo- and regioselective introduction of a variety of functional groups at C1, C2, C3 as well as C4 carbons of the sugar. We have developed a practical approach for stereoselective mono- and diamination of glycals and over the years utilized these scaffolds for the synthesis of a variety of biologically important nitrogen heterocycles and carbocycles through a "Diversity Oriented Approach". Our synthetic strategy in this direction mainly relied on the cleavage of ring O-C bond of the sugar followed by an "intramolecular recombination" reaction. Utilizing this strategy, we have accomplished the synthesis of several biologically important natural products, their analogues and related unnatural derivatives. Examples of such compounds reported from our group include polyhydroxypyrrolidines, DMDP, anisomycin, steviamine, pochonicine, conduramines, bulgecinine, aminocyclitols, azepanes, 4-hydroxy-D-proline, azanucleosides and their analogues. A personal account highlighting these syntheses is presented here.

Cytochrome P450 catalyses the 29-carboxyl group formation of celastrol

Celastrol, a potent anticancer and anti-obesity drug, was first isolated from Tripterygium wilfordii Hook. f. and it is produced in small quantities in many members of the Celastraceae family. The heterologous reconstitution of celastrol biosynthesis could be a promising method for the efficient production of celastrol and natural and unnatural derivatives thereof, yet only part of the biosynthetic pathway is known. Here, we report a cytochrome P450 monooxygenase (TwCYP712K1) from T. wilfordii that performs the three-step oxidation of friedelin to polpunonic acid in the celastrol pathway. Heterologous expression of TwCYP712K1 showed that TwCYP712K1 catalyses not only the transformation of friedelin to polpunonic acid but also the oxidation of β-amyrin or α-amyrin. The role of TwCYP712K1 in the biosynthesis of celastrol was further revealed via RNA interference. Some key residues of TwCYP712K1 were also screened by molecular docking and site-directed mutagenesis. Our results lay a solid foundation for further elucidating the biosynthesis of celastrol and related triterpenoids.

Four-Step Access to the Sesquiterpene Natural Product Presilphiperfolan-1β-ol and Unnatural Derivatives via Supramolecular Catalysis.

Terpenes constitute one of the most structurally varied classes of natural products. A wide range of these structures are produced in nature by type I terpene cyclase enzymes from one single substrate. However, such reactivity has proven difficult to reproduce in solution with man-made systems. Herein we report the shortest synthesis of the tricyclic sesquiterpene presilphiperfolan-1β-ol to date, utilizing the supramolecular resorcinarene capsule as catalyst for the key step. This synthetic approach also allows access to unnatural derivatives of the natural product, which would not be accessible through the biosynthetic machinery. Additionally, this study provides useful insight into the biosynthesis of the presilphiperfolanol natural products, including the first experimental evidence consistent with the proposed biosynthetic connection between caryophyllene and the presilphiperfolanols.

A Versatile Chemoenzymatic Synthesis for the Discovery of Potent Cryptophycin Analogs.

The cryptophycins are a family of macrocyclic depsipeptide natural products that display exceptionally potent antiproliferative activity against drug-resistant cancers. Unique challenges facing the synthesis and derivatization of this complex group of molecules motivated us to investigate a chemoenzymatic synthesis designed to access new analogs for biological evaluation. The cryptophycin thioesterase (CrpTE) and the cryptophycin epoxidase (CrpE) are a versatile set of enzymes that catalyze macrocyclization and epoxidation of over 20 natural cryptophycin metabolites. Thus, we envisioned a drug development strategy involving their use as standalone biocatalysts for production of unnatural derivatives. Herein, we developed a scalable synthesis of 12 new unit A-B-C-D linear chain elongation intermediates containing heterocyclic aromatic groups as alternatives to the native unit A benzyl group. N-Acetyl cysteamine activated forms of each intermediate were assessed for conversion to macrocyclic products using wild type CrpTE, which demonstrated the exceptional flexibility of this enzyme. Semipreparative scale reactions were conducted for isolation and structural characterization of new cryptophycins. Each was then evaluated as a substrate for CrpE P450 and its ability to generate the epoxidized products from these substrates that possess altered electronics at the unit A styrenyl double bond position. Finally, biological evaluation of the new cryptophycins revealed a des-β-epoxy analog with low picomolar potency, previously limited to cryptophycins bearing epoxide functionality.

A Divergent Synthetic Route to the Vallesamidine and Schizozygine Alkaloids: Total Synthesis of (+)-Vallesamidine and (+)-14,15-Dehydrostrempeliopine.

The total synthesis of representative members of the schizozygine alkaloids, (+)-vallesamidine and (+)-14,15-dehydrostrempeliopine, were completed from a late-stage divergent intermediate. The synthesis took advantage of efficient nitro-group reactions with the A/B/C ring skeleton constructed concisely on a gram scale through an asymmetric Michael addition, nitro-Mannich/lactamisation, Tsuji-Trost allylation, and intramolecular C-N coupling reaction. Other key features of the synthesis are a novel [1,4] hydride transfer/Mannich-type cyclisation to build ring E and a diastereoselective ring-closing metathesis reaction to construct ring D. This approach gave access to a late-stage C14,C15 alkene divergent intermediate that could be simply transformed into (+)-vallesamidine, (+)-14,15-dehydrostrempeliopine, and potentially other schizozygine alkaloids and unnatural derivatives.

A Divergent Synthetic Route to the Vallesamidine and Schizozygine Alkaloids: Total Synthesis of (+)‐Vallesamidine and (+)‐14,15‐Dehydrostrempeliopine

AbstractThe total synthesis of representative members of the schizozygine alkaloids, (+)‐vallesamidine and (+)‐14,15‐dehydrostrempeliopine, were completed from a late‐stage divergent intermediate. The synthesis took advantage of efficient nitro‐group reactions with the A/B/C ring skeleton constructed concisely on a gram scale through an asymmetric Michael addition, nitro‐Mannich/lactamisation, Tsuji–Trost allylation, and intramolecular C−N coupling reaction. Other key features of the synthesis are a novel [1,4] hydride transfer/Mannich‐type cyclisation to build ring E and a diastereoselective ring‐closing metathesis reaction to construct ring D. This approach gave access to a late‐stage C14,C15 alkene divergent intermediate that could be simply transformed into (+)‐vallesamidine, (+)‐14,15‐dehydrostrempeliopine, and potentially other schizozygine alkaloids and unnatural derivatives.

Tetrodotoxin: History, Biology, and Synthesis.

This review provides a comprehensive coverage of the history, biology and chemistry of tetrodotoxin (TTX). It traces the origin of this remarkable molecule all the way back to the ancient Chinese medicine records. The discovery of biological activity, isolation, and a brief overview of structure elucidation are summarized. Next, the biology of TTX is discussed, primarily in the context of its activity in the sodium channels, its anesthetic properties, and its potential use in cancer treatment or drug addiction. Biosynthesis of TTX is covered before the discussion of the total syntheses. All total, formal or partial syntheses are covered but those total syntheses that have been discussed in previous reviews are only briefly summarized. Finally, the synthesis of natural and unnatural derivatives is surveyed, and a conclusion and outlook are provided for this very extensive field of endeavor. To the best of our knowledge the literature coverage is complete up to December 2018.

Enantioseparation of tryptophan and its unnatural derivatives by nano-LC on CSP-teicoplanin silica based.

This work deals with the potentiality of nano liquid chromatography (Nano-LC) for the chiral separation of racemic mixture of tryptophan and some selected derivatives by using 100µm i.d. fused silica capillary packed with teicoplanin bonded to 5µm diol silica stationary phase. The experiments were carried out by using a cheap and laboratory-assembled nano-LC-UV system. Elution was done in an isocratic mode using a polar organic mobile phase. In order to find the optimum chiral separation of the studied enantiomers, some chromatographic experimental parameters were systematically studied and optimized. Among them, mobile phase composition, namely organic modifier type and concentration, buffer type and pHand aqueous content and sample solvent dilution on retention time, retention factor and enantioresolution factor were studied. Baseline enantioresolution and good peak shape was achieved utilizing the mobile phase containing 40mM ammonium formate at pHpH2.5 in ACN/water/acetone (60:30:10, v/v/v) at 520nL/min in less than 8min analysis time.

Total synthesis of the linear and angular 3-methylated regioisomers of the marine natural product Kealiiquinone and biological evaluation of related Leucetta sp. alkaloids on human breast cancer

A straightforward, concise, and building block-scalable, seven-step total synthesis of the unnatural 3-methylated regioisomer analog of the marine alkaloid kealiiquinone is described. The construction by a regioselective way of the 4-anisyl-3-methylbenzimidazolone core of this analog, as well as the homologation of the aromatic π-system through a 4π ring opening/6π ring closure sequence for the 1,4-quinoid ring construction by using the squaric acid chemistry, are the key features of this procedure. In light that only one total synthesis of this 3-methyl-iso-kealiiquinone has been described by using a Friedel−Crafts reaction as the key step, herein, we present our pericyclic-based strategy which was successfully concluded with the aforementioned alkaloid synthesis. The biological evaluation of these unnatural derivatives was carried out on MDA-MB231 human cancer cell lines of breast. The cis-platin (CDDP) was used as a positive control of activity.

Reprogramming of the antimycin NRPS-PKS assembly lines inspired by gene evolution

Reprogramming of the NRPS/PKS assembly line is an attractive method for the production of new bioactive molecules. However, it is usually hampered by the loss of intimate domain/module interactions required for the precise control of chain transfer and elongation reactions. In this study, we first establish heterologous expression systems of the unique antimycin-type cyclic depsipeptides: JBIR-06 (tri-lactone) and neoantimycin (tetra-lactone), and engineer their biosyntheses by taking advantage of bioinformatic analyses and evolutionary insights. As a result, we successfully accomplish three manipulations: (i) ring contraction of neoantimycin (from tetra-lactone to tri-lactone), (ii) ring expansion of JBIR-06 (from tri-lactone to tetra-lactone), and (iii) alkyl chain diversification of JBIR-06 by the incorporation of various alkylmalonyl-CoA extender units, to generate a set of unnatural derivatives in practical yields. This study presents a useful strategy for engineering NRPS-PKS module enzymes, based on nature’s diversification of the domain and module organizations.

Site‐Selective C–H Bond Activation/Functionalization of Alpha‐Amino Acids and Peptide‐Like Derivatives

Amino acids and peptides play an important role in nature, as well as in organic and pharmaceutical chemistry. Therefore, the easy, straightforward and versatile access of biogenic and unnatural derivatives is still highly demanding. This micro‐review intends to provide to the reader the current state of the art on site‐selective C–H bond functionalization technology applied on α‐amino acids and peptides, focusing on the different C–H positions that can successfully be addressed to date. It is structured in two main parts implying i) the α‐C–H functionalization at the peptide backbone and ii) the modification of the side‐chain. Herein, metal‐catalyzed C–H activation, oxidative C–H functionalization, as well as light mediated approaches, are discussed considering the challenges in reactivity and selectivity. The great potential of these transformations is also depicted with the derivatization and macrocyclization of complex peptides leading to products of pharmaceutical importance.