Cyclization Efficiency Research Articles

A Reusable Polystyrene Support for Sustainable yet Cost-effective Octreotide Synthesis.

The synthesis of peptide drugs has become facile with the use of supports that enable easy separation of the growing peptide. Peptide synthesis on insoluble supports typically employs excess reagents to enhance reaction efficiency and faces challenges during intramolecular cyclization. A non-crosslinked soluble polystyrene support is reported herein that improves cyclization efficiency on the support by using spacers. The support efficiency is illustrated by synthesizing octreotide drug in a scalable manner by on-support cyclization. The methodology drastically reduces the reagent waste, raw-material cost, and solvent requirement. The support can also be recovered and re-used up to 3 times making this a very sustainable method.

Unique Electron Donor–Acceptor Complex Conformation Ensures Both the Efficiency and Enantioselectivity of Photoinduced Radical Cyclization in a Non-natural Photoenzyme

Unique Electron Donor–Acceptor Complex Conformation Ensures Both the Efficiency and Enantioselectivity of Photoinduced Radical Cyclization in a Non-natural Photoenzyme

Frustrated Lewis pairs on facet-engineered CeO2 boost the conversion of low-concentration CO2 into cyclic ureas with assistance of diamine platforms

The development of efficient processes for value-added utilization of low-concentration CO2 is an on-going challenge. In this work, we present a sequential system based on diamine platforms for the direct utilization of low-concentration sources of CO2 (≤15%) in the production of cyclic ureas. Specifically, the low-concentration CO2 is captured by ethylenediamine to form ethylenediamine carbamate (EDA-CA), which subsequently undergoes the intramolecular dehydration to give ethyleneurea (EU) in the presence of frustrated Lewis pairs (FLPs) on the facet-engineered CeO2. Remarkably, the productivity of EU obtained from EDA-CA in the sequential system (9.5 mmol·gcat−1·h−1) is ∼4 times higher compared to the traditional catalytic system (2.4 mmol·gcat−1·h−1) using ethylenediamine and pure CO2 (3 MPa). Density functional theory calculations demonstrated that the FLPs on facet-engineered CeO2 significantly reduce the energy barrier for the nucleophilic attack of N-containing segment towards the carbonyl carbon in EDA-CA (rate-limiting step), ultimately optimizing the intramolecular cyclization efficiency of EDA-CA. This work not only provides an innovation tandem approach that enables the production of cyclic ureas from low-concentration CO2, but also opens up a promising avenue for the direct utilization of low-concentration CO2 for value-added applications in the future.

Influence of polymer amphiphilicity on the cyclization efficiency of poly(2-oxazoline)s

The steadily increasing interest in cyclic polymers has led to a plethora of proposed applications, particularly as biomaterials. However, the large-scale synthesis of cyclic polymers remains one of the greatest challenges in the field. One reason for this is the poorly understood role of the polymer structure on the cyclization efficiency, leading to synthesis protocols that must be optimized on a case-by-case basis without consideration for the chemical nature of the polymer. Recently, we found through a combination of laboratory experiments and molecular dynamics simulations that the polymer backbone flexibility has a major influence on the cyclization efficiency of poly(cyclic imino ether)s. Concurrently, we identified that a more flexible polymer backbone was also better solvated in the given organic solvent. To deconvolute the importance of the polymer backbone flexibility and solvation, in this study a variety of α-alkyne, ω-azide terminated (co)polymers of 2-ethyl-2-oxazoline and 2–n-butyl-2-oxazoline with different amphiphilicities, but the same polymer backbone, have been synthesized and subjected to copper-catalysed azide-alkyne cycloaddition (CuAAC) reactions in water. In agreement with our previous studies, it was observed that an increase in polymer chain hydrophobicity, and hence a decrease in solvation, leads to a significant decrease in cyclization efficiency. To further understand the effect of different CuAAC reaction conditions on the reaction outcome, different polymer concentrations and reaction temperatures were screened. This study highlights the importance of polymer amphiphilicity, and more generally solvation, in CuAAC protocols aimed towards the synthesis of cyclic polymers.

Vaccines' New Era-RNA Vaccine.

RNA vaccines, including conventional messenger RNA (mRNA) vaccines, circular RNA (circRNA) vaccines, and self-amplifying RNA (saRNA) vaccines, have ushered in a promising future and revolutionized vaccine development. The success of mRNA vaccines in combating the COVID-19 pandemic caused by the SARS-CoV-2 virus that emerged in 2019 has highlighted the potential of RNA vaccines. These vaccines possess several advantages, such as high efficacy, adaptability, simplicity in antigen design, and the ability to induce both humoral and cellular immunity. They also offer rapid and cost-effective manufacturing, flexibility to target emerging or mutant pathogens and a potential approach for clearing immunotolerant microbes by targeting bacterial or parasitic survival mechanisms. The self-adjuvant effect of mRNA-lipid nanoparticle (LNP) formulations or circular RNA further enhances the potential of RNA vaccines. However, some challenges need to be addressed. These include the technology's immaturity, high research expenses, limited duration of antibody response, mRNA instability, low efficiency of circRNA cyclization, and the production of double-stranded RNA as a side product. These factors hinder the widespread adoption and utilization of RNA vaccines, particularly in developing countries. This review provides a comprehensive overview of mRNA, circRNA, and saRNA vaccines for infectious diseases while also discussing their development, current applications, and challenges.

A molecular dynamics perspective on the cyclization efficiency for poly(2-oxazoline)s and poly(2-oxazine)s

Poly(2-n-propyl-2-oxazine) is better solvated and shows higher backbone flexibility than its oxazoline analogue in dichloromethane, resulting in short distances between chain ends and ultimately increased cyclization efficiency.

Double crossing conical intersections and anti-Vavilov fluorescence in tetraphenyl ethylene.

Conical intersections (CIs) provide effective fast nonradiative decay pathways for electronic excitation, which can significantly influence molecular photoluminescence properties. However, in many cases, crossing a CI does not have direct observables, making studies of CIs experimentally challenging. Herein, the theoretically predicted double CIs by cis-trans twisting and cyclization in tetraphenyl ethylene, a well-known aggregation-induced emission molecule, are investigated with excitation dependent ultrafast UV/IR spectroscopy and fluorescence. Both the fluorescence quantum yield and the efficiency of cyclization are found to be smaller with a shorter excitation wavelength. An abrupt change occurs at about 300-310nm. The results imply that crossing the twisting CI has a larger barrier than the cyclization CI, and the cis-trans twisting motion is probably involved with large solvation reorganization.

Site-Specific Protein Modifications by an Engineered Asparaginyl Endopeptidase from Viola canadensis.

Asparaginyl endopeptidases (AEPs) or legumains are Asn/Asp (Asx)-specific proteases that break peptide bonds, but also function as peptide asparaginyl ligases (PALs) that make peptide bonds. This ligase activity can be used for site-specific protein modifications in biochemical and biotechnological applications. Although AEPs are common, PALs are rare. We previously proposed ligase activity determinants (LADs) of these enzymes that could determine whether they catalyze formation or breakage of peptide bonds. LADs are key residues forming the S2 and S1′ substrate-binding pockets flanking the S1 active site. Here, we build on the LAD hypothesis with the engineering of ligases from proteases by mutating the S2 and S1′ pockets of VcAEP, an AEP from Viola canadensis. Wild type VcAEP yields <5% cyclic product from a linear substrate at pH 6.5, whereas the single mutants VcAEP-V238A (Vc1a) and VcAEP-Y168A (Vc1b) targeting the S2 and S1′ substrate-binding pockets yielded 34 and 61% cyclic products, respectively. The double mutant VcAEP-V238A/Y168A (Vc1c) targeting both the S2 and S1′ substrate-binding pockets yielded >90% cyclic products. Vc1c had cyclization efficiency of 917,759 M−1s−1, which is one of the fastest rates for ligases yet reported. Vc1c is useful for protein engineering applications, including labeling of DARPins and cell surface MCF-7, as well as producing cyclic protein sfGFP. Together, our work validates the importance of LADs for AEP ligase activity and provides valuable tools for site-specific modification of proteins and biologics.

Effects of protecting group and counter‐anion on fluorination, bromination, and intramolecular cyclization of phenethylamine diaryliodonium salts: Quantum chemical analysis

AbstractWe analyze the effects of protecting group(s) and counter‐anion on the relative efficiency of fluorination, bromination and intramolecular cyclization of phenethylamine diaryliodonium salts (1) for nucleophilic fluorination using CsF as a model system for the synthesis of radiopharmaceuticals such as 18F‐dopa and 18F‐dopamine. We demonstrate by quantum chemical analysis that protection of –NH2 by –Boc groups strongly influences the relative yields of nucleophilic fluorination versus side reactions (bromination and intramolecular cyclization) through intricate interactions with iodonium, counter‐anion (Br−, F−, or OMs−) to iodonium, and counter‐cation Cs+ to the nucleophile F−. Protection of the amino by one or two –Boc group(s) render fluorination to be strongly favored over bromination/cyclization. Possibility of direct attack by counter‐anion is assessed. We propose that mesylate (–OMs−) would be much better counter‐anion than Br− because of its much weaker nucleophilicity. F− is also preferred as a counter‐anion, because using it would exclusively give fluorinated product. We also estimate the capability of crown ether (18‐crown‐6) as a Lewis base promoter for enhancing the rates of fluorination.

Gas‐Phase Transformation of Fluorinated Benzoporphyrins to Porphyrin‐Embedded Conical Nanocarbons

Geodesic nitrogen‐containing graphene fragments are interesting candidates for various material applications, but the available synthetic protocols, which need to overcome intrinsic strain energy during the formation of the bowl‐shaped skeletons, are often incompatible with heteroatom‐embedded structures. Through this mass spectrometry‐based gas‐phase study, we show by means of collision‐induced dissociation experiments and supported by density functional theory calculations, the first evidence for the formation of a porphyrin‐embedded conical nanocarbon. The influences of metalation and functionalization of the used tetrabenzoporphyrins have been investigated, which revealed different cyclization efficiencies, different ionization possibilities, and a variation of the dissociation pathway. Our results suggest a stepwise process for HF elimination from the fjord region, which supports a selective pathway towards bent nitrogen‐containing graphene fragments.

Enantioselective, Aerobic Copper-Catalyzed Intramolecular Carboamination and Carboetherification of Unactivated Alkenes.

Reduction of waste is an important goal of modern organic synthesis. We report herein oxidase reactivity for enantioselective intramolecular copper-catalyzed alkene carboamination and carboetherification reactions where previously used stoichiometric MnO2 has been replaced with oxygen. This substitution was risky as the reaction mechanism is thought to involve C-C bond formation via addition of alkyl carbon radicals to arenes. Such intermediates are also susceptible to C-O bond formation via O2 addition. Control of absolute stereochemistry under aerobic conditions was also uncertain. The oxidative cyclization efficiencies appear to track with the ease of the radical addition to the arenes.

Construction of Highly Emissive Pt(II) Metallacycles upon Irradiation

Summary of main observation and conclusionPhotoswitchable or photoactivatable fluorescent species have been found wide applications within supramolecular chemistry and materials science. In this study, we successfully constructed two highly emissive Pt(II) metallacycles from the diarylethene ligands via coordination‐driven self‐assembly. Different from the most known fluorescent metallacycles, the obtained metallacycles have displayed “turn‐on” fluorescence switching. They are non‐fluorescent in solution, but they emit highly yellow or orange fluorescence under ultraviolet irradiation. The metallacycles were well characterized by 1H NMR, 31P NMR and ESI‐TOF‐MS. The photochromic properties of the resultant metallacycles were investigated by 1H NMR, 31P NMR, UV/Vis spectrum and fluorescence spectrum. Notably, NMR studies revealed that these two metallacycles featured excellent cyclization efficiency (90% conversion efficiency). Moreover, the closed‐ring isomers of the metallacycles displayed relatively high quantum yield (ΦF = 0.5). DFT simulations demonstrated that the antiparallel configuration of the diarylethene ligand had an angle closed to 120°, which was more stable in energy compared to the parallel configuration, thus allowing for the facile construction of highly emissive metallacycles. We believe that such highly emissive metallacycles which are in‐situ prepared upon irradiation can be used as new fluorescence materials for sensing and bioimaging in the future.

Measurement of the Length Dependence of DNA Cyclization using Next Generation Sequencing

DNA flexibility is important both for fundamental biophysics and also because DNA flexibility affects DNA packaging and the regulation of gene expression through DNA looping. Historically DNA flexibility has been studied with experiments ranging from biochemical ring closure or DNA looping experiments to AFM, crystallography, and tethered particle microscopy. Even so, the flexibility of DNA in vitro and in vivo remains controversial. We have constructed a library of DNA molecules ranging from 125 to 225 base pairs, via ligation of pools of synthetic DNA of different lengths and PCR, yielding 1023 distinct sequences. The design incorporated barcoding for redundant identification of each molecule, allowing for a ligation reaction to be performed on the entire library in the same reaction mixture. Two different DNA concentrations were used to promote either unimolecular cyclization or bimolecular ligation and thereby explore a wide range of cyclization efficiencies (J factors). A portion of each reaction mixture was treated with BAL-31 to destroy non-cyclized molecules. All products were linearized by restriction digestion and Illumina indices were added to the four mixtures of cyclized and non-cyclized products. The initial library and all of the reaction mixtures were sequenced in a single Illumina MiSeq 300 base pair paired end run. From the roughly 30 million sequence reads obtained, we expect to extract J factors for each of the 1023 molecules in the library, giving an overlapping set of J factors as a function of length. The analysis of the length dependence of ring closure will be presented. We believe this data set will be valuable for improving course grain modeling of DNA. The same methodology should be applicable to DNA loops anchored by proteins.

Investigation for the cyclization efficiency of linear tetrapeptides: Synthesis of tentoxin B and dihydrotentoxin

Investigation of the cyclization efficiency of N-methyl linear tetrapeptides using a molecular modeling study and chemical synthesis is described. The linear peptide with two N-methyl groups, MeAla-Leu-MePhe-Gly, forms γ-turn like conformation with the amine at N-terminus and the carbonyl at C-terminus in closer proximity to give the desired cyclic tetrapeptide, dihydrotentoxin. In addition, synthesis of tentoxin B by the combination of Fmoc solid-phase peptide synthesis and cyclization in solution phase has been reported. An unusual amino acid, an L-N-methyl-β-hydroxyphenylalanine derivative, which was assembled on solid support, was prepared from ethyl cinnamate. Cyclic tetrapeptide formation and cleavage of benzyl ether were optimized with DIPCI/HOBt/DIPEA and Et3SiH/Pd(OH)2, respectively.

Intein-mediated backbone cyclization of entolimod confers enhanced radioprotective activity in mouse models.

BackgroundEntolimod is a Salmonella enterica flagellin derivate. Previous work has demonstrated that entolimod effectively protects mice and non-human primates from ionizing radiation. However, it caused a “flu-like” syndrome after radioprotective and anticancer clinical application, indicating some type of immunogenicity and toxicity. Cyclization is commonly used to improve the in vivo stability and activity of peptides and proteins.MethodsWe designed and constructed cyclic entolimod using split Nostoc punctiforme DnaE intein with almost 100% cyclization efficiency. We adopted different strategies to purify the linear and circular entolimod due to their different topologies. Both of linear and circular entolimod were first purified by Ni-chelating affinity chromatography, and then the linear and circular entolimod were purified by size-exclusion and ion-exchange chromatography, respectively.ResultsThe circular entolimod showed significantly increased both the in vitro NF-κB signaling and in vivo radioprotective activity in mice.ConclusionOur data indicates that circular entolimod might be a good candidate for further clinical investigation.

Precursor Manipulation in Glycopeptide Antibiotic Biosynthesis: Are β-Amino Acids Compatible with the Oxidative Cyclization Cascade?

Natural products such as the glycopeptide antibiotics (GPAs, including vancomycin and teicoplanin) are of great pharmaceutical importance due to their use against Gram-positive bacteria such as methicillin-resistant Staphylococcus aureus. GPAs are assembled in a complex process based on nonribosomal peptide synthesis and late-stage, multistep cross-linking of the linear heptapeptide performed by cytochrome P450 monooxygenases. These P450 enzymes demonstrate varying degrees of substrate selectivity toward the linear peptide precursor, with limited information available about their tolerance regarding modifications to amino acid residues within the essential antibiotic core of the GPA. In order to test the acceptance of altered residues by the P450-catalyzed cyclization cascade, we have explored the use of β-amino acids in both variable and highly conserved positions within GPA peptides. Our results indicate that the incorporation of β-amino acids at the C-terminus of the peptide leads to a dramatic reduction in the efficiency of peptide cyclization by the P450s during GPA biosynthesis, whereas replacement of residue 3 is well tolerated by the same enzymes. These results show that maintaining the C-terminal 3,5-dihydroxyphenylglycine residue is of key importance to maintain the efficiency of this complex and essential enzymatic cross-linking process.

Head-to-tail macrocyclization of cysteine-free peptides using an o-aminoanilide linker

A head-to-tail macrocyclization protocol for the preparation of cysteine-free cyclic peptides was investigated. The o-aminoanilide linker constructed in the peptide sequence by a standard Fmoc-based peptide synthesis procedure was subjected to nitrite-mediated activation under acidic conditions toward N-acyl benzotriazole as the active ester species. The subsequent cyclization smoothly proceeded by neutralization in the presence of additives such as 1-hydroxybenzotriazole (HOBt) and 1-hydroxy-7-azabenzotriazole (HOAt) to afford the expected cyclic pentapeptide, a CXCR4 antagonist. The cyclization efficiencies were dependent on the precursor open-chain sequence. The application of this step-wise activation-cyclization protocol to microflow reaction systems is also described.

Synthesis of Cyclic Peptides as Potential Anti-Malarials.

The results from the synthesis of peptides by Fmoc/SPPS on a 2-CTC resin and then lactamization in solution or solid phase for the preparation of cyclopeptides are presented. Both procedures allow the synthesis of the desired compounds in good to very good yield and with high cyclization efficiency for on-resin macrocyclization. In addition, the activities of the corresponding cyclopeptides against the chloroquine-resistant K1 strain of Plasmodium falciparum were evaluated. Cyclo-Cys(Trt)-Gly-Thr( tBu)-Gly-Cys(Trt)-Gly showed potent in vitro and selective activity against this parasite, EC50 = 28 nM.

Iminoboronate-Mediated Peptide Cyclization with Lysine Homologues

Cyclic peptides are attracting attention of medicinal chemists due to their increased stability in biological milieu as well as improved target binding affinities. Our laboratory has recently reported a powerful cyclization strategy that takes advantage of the spontaneous and reversible conjugation of lysine and a designed amino acid AB3 to give iminoboronates. Herein we report that Dap, a short chain homologue of lysine, displays significantly higher propensity to form imino­boronates and consequently improves the efficiency of peptide cyclization. Importantly, the preferential conjugation of AB3 to Dap allows a facile synthesis of cyclic peptides with free lysine residues.

Seven-Membered Ring-Forming Cyclopolymerization of 1,8-Nonadiyne Derivatives Using Grubbs Catalysts: Rational Design of Monomers and Insights into the Mechanism for Olefin Metathesis Polymerizations

Studies into the cyclopolymerization (CP) of diyne derivatives using metal carbenes have focused on the formation of five- and six-membered rings because these small rings can be easily synthesized while the preparation of medium-sized seven-membered rings are more difficult. For the first time, we achieved the CP forming challenging seven-membered rings as repeat units using Grubbs catalysts by novel design of 1,8-nonadiyne monomers. The key to the successful CP was the introduction of the appropriate aminal and acetal groups, which have short C–N and C–O bonds, and low rotational barriers, thus greatly enhancing the cyclization efficiency. During our mechanistic investigation, we directly observed an actual 14-electron Ru propagating carbene by 1H NMR spectroscopy for the first time during olefin metathesis reaction, presumably because the great steric hindrance from the propagating carbene containing a larger seven-membered ring than five- or six-membered ring retarded the coordination of ligands. We also observed decomposition of the catalysts to ruthenium hydrides during polymerization for the first time. Kinetic studies revealed three interesting features of this 1,8-nonadiyne CP: (i) in contrast to conventional polymerizations, the rate-determining step for the CP of 1,8-nonadiynes was the cyclization step; (ii) the intrinsic reactivity of the acetal monomers was higher than that of the aminal monomers; but (iii) the overall polymerization efficiency of the aminal monomers was higher than that of the acetal monomers because of the higher stability of their carbenes. Finally, we achieved a controlled CP of the aminal monomers using a fast-initiating third-generation Grubbs catalyst. This allowed the synthesis of not only the diblock copolymer containing five- and seven-membered rings but also the triblock copolymer containing five-, six-, and seven-membered rings.