| 32824227 |
Involvement of Reactive Oxygen Species in the Hepatorenal Toxicity of Actinomycin V In Vitro and In Vivo |
10.3390/md18080428. |
Mar Drugs |
Involvement of Reactive Oxygen Species in the Hepatorenal Toxicity of Actinomycin V In Vitro and In Vivo
Abstract
- The high toxicity of actinomycin D (Act D) severely limits its use as a first-line chemotherapeutic agent in the clinic. Actinomycin V (Act V), an analog of Act D, exhibited strong anticancer activity in our previous studies. Here, we provide evidence that Act V has less hepatorenal toxicity than Act D in vitro and in vivo, associated with the reactive oxygen species (ROS) pathway. Compared to Act D, Act V exhibited considerably stronger sensitivity for cancer cells and less toxicity to human normal liver LO-2 and human embryonic kidney 293T cells using the MTT (3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide) assay. Notably, Act V caused less damage to both the liver and kidney than Act D in vivo, indicated by organ to body weight ratios, as well as alanine aminotransferase (ALT), aspartate aminotransferase (AST), and serum creatinine (Scr) levels. Further experiments showed that the ROS pathway is involved in Act V-induced hepatorenal toxicity. Act V generates ROS and accumulates malondialdehyde (MDA), reducing levels of superoxide dismutase (SOD) and glutathione (GSH) in LO-2 and 293T cells. These findings indicate that Act V induces less hepatorenal toxicity than Act D in vitro and in vivo and merits further development as a potential therapeutic agent for the treatment of cancer.
|
| 32840352 |
Structure-Based Design of VapC-Activating Stapled Peptides |
10.1021/acschembio.0c00492. |
ACS Chem Biol |
Structure-Based Design of VapC-Activating Stapled Peptides
Abstract
- Toxin-antitoxin (TA) systems have been considered essential factors for bacterial survival. During our drug development program aimed against tuberculosis (TB), we discovered certain peptides that mimic the binding of the VapBC30 complex, leading to the arrest of bacterial cell growth and eventually cell death. Herein, we optimized these candidate peptides based on a hydrocarbon stapling strategy and performed biological evaluations. The peptide successfully penetrated cell membranes and exerted bactericidal activity at a minimum inhibitory concentration that inhibited 50% of the isolates (MIC) < 6.25 μM. With the aid of structural and biochemical information for the VapBC30 TA system from , we suggest potential antimicrobial agents that could provide a platform to establish a novel antibacterial strategy. Reflecting the limited number of therapeutic agents targeting TA systems, we believe that this study not only provides chemical tools for exploring the biological events relevant to TA systems but also opens a new gateway toward TB drug discovery.
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| 33002682 |
Ahp-Cyclodepsipeptides as tunable inhibitors of human neutrophil elastase and kallikrein 7: Total synthesis of tutuilamide A, serine protease selectivity profile and comparison with lyngbyastatin 7 |
10.1016/j.bmc.2020.115756. |
Bioorg Med Chem |
Ahp-Cyclodepsipeptides as tunable inhibitors of human neutrophil elastase and kallikrein 7: Total synthesis of tutuilamide A, serine protease selectivity profile and comparison with lyngbyastatin 7
Abstract
- We describe the total synthesis of tutuilamide A, a potent porcine pancreatic elastase (PPE) inhibitor and a representative member of the 3-amino-6-hydroxy-2-piperidone (Ahp) cyclodepsipeptide family, isolated from marine cyanobacteria. The Ahp unit serves as a pharmacophore and the adjacent 2-amino-2-butenoic acid (Abu) is a main driver of the selectivity among serine proteases. We adapted our previous convergent strategy to generate the macrocycle, common with lyngbyastatin 7 and related elastase inhibitors, and then appended the tutuilamide A-specific side chain bearing a vinyl chloride. Tutuilamide A and lyngbyastatin 7 were evaluated side by side for the inhibition of the disease-relevant human neutrophil elastase (HNE). Tutuilamide A and lyngbyastatin 7 were approximately equipotent against HNE, while tutuilamide A was previously shown to be more active against PPE compared with lyngbyastatin 7, further demonstrating that the side chain provides opportunities to not only modulate potency but also selectivity among proteases of the same function from different organisms. Profiling of tutuilamide A against mainly human serine proteases revealed high selectivity for HNE (IC 0.73 nM) and pleiotropic activity against kallikrein 7 (KLK7, IC 5.0 nM), without affecting other kallikreins, similarly to lyngbyastatin 7 (IC 0.85 nM for HNE and 3.1 nM for KLK7). A comprehensive molecular docking study for elastases and KLK7 afforded deeper insight into the intricate differences between inhibitor interactions with HNE and PPE, accounting for the differential activities for both compounds. The synthesis and molecular studies serve as a proof-of-concept that the macrocyclic scaffold can be diversified to fine-tune the activity of serine protease inhibitors.
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| 33019526 |
Identification of Conomarphin Variants in the Venom and the Effect of Sequence and PTM Variations on Conomarphin Conformations |
10.3390/md18100503. |
Mar Drugs |
Identification of Conomarphin Variants in the Venom and the Effect of Sequence and PTM Variations on Conomarphin Conformations
Abstract
- Marine cone snails belonging to the Conidae family make use of neuroactive peptides in their venom to capture prey. Here we report the proteome profile of the venom duct of , a cone snail belonging to the Tesseliconus clade. Through tandem mass spectrometry and database searching against the transcriptome and the ConoServer database, we identified 24 unique conopeptide sequences in the venom duct. The majority of these peptides belong to the T and M gene superfamilies and are disulfide-bonded, with cysteine frameworks V, XIV, VI/VII, and III being the most abundant. All seven of the Cys-free peptides are conomarphin variants belonging to the M superfamily that eluted out as dominant peaks in the chromatogram. These conomarphins vary not only in amino acid residues in select positions along the backbone but also have one or more post-translational modifications (PTMs) such as proline hydroxylation, C-term amidation, and γ-carboxylation of glutamic acid. Using molecular dynamics simulations, the conomarphin variants were predicted to predominantly have hairpin-like or elongated structures in acidic pH. These two structures were found to have significant differences in electrostatic properties and the inclusion of PTMs seems to complement this disparity. The presence of polar PTMs (hydroxyproline and γ-carboxyglutamic acid) also appear to stabilize hydrogen bond networks in these conformations. Furthermore, these predicted structures are pH sensitive, becoming more spherical and compact at higher pH. The subtle conformational variations observed here might play an important role in the selection and binding of the peptides to their molecular targets.
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| 33284578 |
Unnarmicin D, an Anti-inflammatory Cyanobacterial Metabolite with δ and μ Opioid Binding Activity Discovered via a Pipeline Approach Designed to Target Neurotherapeutics |
10.1021/acschemneuro.0c00686. |
ACS Chem Neurosci |
Unnarmicin D, an Anti-inflammatory Cyanobacterial Metabolite with δ and μ Opioid Binding Activity Discovered via a Pipeline Approach Designed to Target Neurotherapeutics
Abstract
- To combat the bottlenecks in drug discovery and development, a pipeline to identify neuropharmacological candidates using , , and receptor specific assays was devised. The focus of this pipeline was to identify metabolites with the ability to reduce neuroinflammation, due to the implications that chronic neuroinflammation has in chronic pain and neurodegenerative diseases. A library of pure compounds isolated from the cyanobacterium was evaluated using this method. analysis of drug likelihood and permeability analysis using the parallel artificial membrane permeability assay (PAMPA) highlighted multiple metabolites of interest from the library. Murine BV-2 microglia were used in conjunction with the Griess assay to determine if metabolites could reduce lipopolysaccharide induced neuroinflammation followed by analysis of pro-inflammatory cytokine concentrations in the supernatant of the treated cell cultures. The nontoxic metabolite unnarmicin D was further evaluated due to its moderate permeability in the PAMPA assay, promising ADME data, modulation of all cytokines tested, and prediction as an opioid receptor ligand. Molecular modeling of unnarmicin D to the μ and δ opioid receptors showed strong theoretical binding potential to the μ opioid receptor. binding assays validated this pipeline showing low micromolar binding affinity for the μ opioid receptor launching the potential for further analysis of unnarmicin D derivatives for the treatment of pain and neuroinflammation related diseases.
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| 33287430 |
MALDI MSI Reveals the Spatial Distribution of Protein Markers in Tracheobronchial Lymph Nodes and Lung of Pigs after Respiratory Infection |
10.3390/molecules25235723. |
Molecules |
MALDI MSI Reveals the Spatial Distribution of Protein Markers in Tracheobronchial Lymph Nodes and Lung of Pigs after Respiratory Infection
Abstract
- Respiratory infections are a real threat for humans, and therefore the pig model is of interest for studies. As one of a case for studies, (APP) caused infections and still worries many pig breeders around the world. To better understand the influence of pathogenic effect of APP on a respiratory system-lungs and tracheobronchial lymph nodes (TBLN), we aimed to employ matrix-assisted laser desorption/ionization time-of-flight mass spectrometry imaging (MALDI-TOF MSI). In this study, six pigs were intranasally infected by APP and two were used as non-infected control, and 48 cryosections have been obtained. MALDI-TOF MSI and immunohistochemistry (IHC) were used to study spatial distribution of infectious markers, especially interleukins, in cryosections of porcine tissues of lungs (necrotic area, marginal zone) and tracheobronchial lymph nodes (TBLN) from pigs infected by APP. CD163, interleukin 1β (IL‑1β) and a protegrin-4 precursor were successfully detected based on their tryptic fragments. CD163 and IL‑1β were confirmed also by IHC. The protegrin-4 precursor was identified by MALDI-TOF/TOF directly on the tissue cryosections. CD163, IL‑1β and protegrin‑4 precursor were all significantly ( < 0.001) more expressed in necrotic areas of lungs infected by APP than in marginal zone, TBLN and in control lungs.
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| 33296204 |
Evaluation of the Aphrodisiac Activity of a Cyclotide Extract from |
10.1021/acs.jnatprod.0c01045. |
J Nat Prod |
Evaluation of the Aphrodisiac Activity of a Cyclotide Extract from
Abstract
- is an Indian folk medicinal herb that has been widely used as a libido enhancer. This plant belongs to the Violaceae plant family, which ubiquitously contains disulfide-rich cyclic peptides named cyclotides. Cyclotides are an expanding plant-derived peptide family with numerous interesting bioactivities, and their unusual stability against proteolysis has attracted much attention in drug design applications. Recently, has been reported to be a rich source of cyclotides, and hence, it was of interest to investigate whether cyclotides contribute to its aphrodisiac activity. In this study, we evaluated the aphrodisiac activity of the herbal powder, extract, and the most abundant cyclotide, hyen D, extracted from on rats in a single dose regimen. After dosing, the sexual behaviors of male rats were observed, recorded, analyzed, and compared with those of the vehicle group. The results show that the extract and hyen D significantly decreased the intromission latency of sexually naïve male rats and the extract improved a range of other measured sexual parameters. The results suggest that the extract could enhance libido as well as facilitate erectile function in male rats and that the cyclotide hyen D could contribute to the libido-enhancing activity of this ethnomedicinal herb.
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| 33310780 |
Stapled Peptides Based on Human Angiotensin-Converting Enzyme 2 (ACE2) Potently Inhibit SARS-CoV-2 Infection |
10.1128/mBio.02451-20. |
mBio |
Stapled Peptides Based on Human Angiotensin-Converting Enzyme 2 (ACE2) Potently Inhibit SARS-CoV-2 Infection
Abstract
- SARS-CoV-2 uses human angiotensin-converting enzyme 2 (ACE2) as the primary receptor to enter host cells and initiate the infection. The critical binding region of ACE2 is an ∼30-amino-acid (aa)-long helix. Here, we report the design of four stapled peptides based on the ACE2 helix, which is expected to bind to SARS-CoV-2 and prevent the binding of the virus to the ACE2 receptor and disrupt the infection. All stapled peptides showed high helical contents (50 to 94% helicity). In contrast, the linear control peptide NYBSP-C showed no helicity (19%). We have evaluated the peptides in a pseudovirus-based single-cycle assay in HT1080/ACE2 cells and human lung cell line A549/ACE2, overexpressing ACE2. Three of the four stapled peptides showed potent antiviral activity in HT1080/ACE2 (50% inhibitory concentration [IC]: 1.9 to 4.1 μM) and A549/ACE2 (IC: 2.2 to 2.8 μM) cells. The linear peptide NYBSP-C and the double-stapled peptide StRIP16, used as controls, showed no antiviral activity. Most significantly, of the stapled peptides show any cytotoxicity at the highest dose tested. We also evaluated the antiviral activity of the peptides by infecting Vero E6 cells with the replication-competent authentic SARS-CoV-2 (US_WA-1/2020). NYBSP-1 was the most efficient, preventing the complete formation of cytopathic effects (CPEs) at an IC of 17.2 μM. NYBSP-2 and NYBSP-4 also prevented the formation of the virus-induced CPE with an IC of about 33 μM. We determined the proteolytic stability of one of the most active stapled peptides, NYBSP-4, in human plasma, which showed a half-life () of >289 min. SARS-CoV-2 is a novel virus with many unknowns. No vaccine or specific therapy is available yet to prevent and treat this deadly virus. Therefore, there is an urgent need to develop novel therapeutics. Structural studies revealed critical interactions between the binding site helix of the ACE2 receptor and SARS-CoV-2 receptor-binding domain (RBD). Therefore, targeting the entry pathway of SARS-CoV-2 is ideal for both prevention and treatment as it blocks the first step of the viral life cycle. We report the design of four double-stapled peptides, three of which showed potent antiviral activity in HT1080/ACE2 cells and human lung carcinoma cells, A549/ACE2. Most significantly, the active stapled peptides with antiviral activity against SARS-CoV-2 showed high α-helicity (60 to 94%). The most active stapled peptide, NYBSP-4, showed substantial resistance to degradation by proteolytic enzymes in human plasma. The lead stapled peptides are expected to pave the way for further optimization of a clinical candidate.
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| 33348729 |
Improved Stability and Activity of a Marine Peptide-N6NH2 against and Its Preliminary Application in Fish |
10.3390/md18120650. |
Mar Drugs |
Improved Stability and Activity of a Marine Peptide-N6NH2 against and Its Preliminary Application in Fish
Abstract
- can cause fatal gastro-/extraintestinal diseases in fish and humans. Overuse of antibiotics has led to antibiotic resistance and contamination in the environment, which highlights the need to find new antimicrobial agents. In this study, the marine peptide-N6 was amidated at its C-terminus to generate N6NH2. The antibacterial activity of N6 and N6NH2 against was evaluated in vitro and in vivo; their stability, toxicity and mode of action were also determined. Minimal inhibitory concentrations (MICs) of N6 and N6NH2 against were 1.29-3.2 μM. Both N6 and N6NH2 killed bacteria by destroying the cell membrane of and binding to lipopolysaccharide (LPS) and genomic DNA. In contrast with N6, N6NH2 improved the stability toward trypsin, reduced hemolysis (by 0.19% at a concentration of 256 μg/mL) and enhanced the ability to penetrate the bacterial outer and inner membrane. In the model of fish peritonitis caused by , superior to norfloxacin, N6NH2 improved the survival rate of fish, reduced the bacterial load on the organs, alleviated the organ injury and regulated the immunity of the liver and kidney. These data suggest that the marine peptide N6NH2 may be a candidate for novel antimicrobial agents against infections.
|
| 33363118 |
Design, Synthesis, and Antitumor Activities Study of Stapled A4K14-Citropin 1.1 Peptides |
10.3389/fchem.2020.616147. |
Front Chem |
Design, Synthesis, and Antitumor Activities Study of Stapled A4K14-Citropin 1.1 Peptides
Abstract
- A4K14-citropin 1.1 is a structurally optimized derivative derived from amphibians' skin secreta peptide Citropin, which exhibits broad biological activities. However, the application of A4K14-citropin 1.1 as a cancer therapeutic is restricted by its structural flexibility. In this study, a series of all-hydrocarbon stapled peptides derivatives of A4K14-citropin 1.1 were designed and synthesized, and their chemical and biological characteristics were also investigated. Among them, A4K14-citropin 1.1-Sp1 and A4K14-citropin 1.1-Sp4 displayed improved helicity levels, greater protease stability, and increased antitumor activity compared with the original peptide, which establishes them as promising lead compounds for novel cancer therapeutics development. These results revealed the important influence of all-hydrocarbon stapling side chain on the secondary structure, hydrolase stability, and biological activity of A4K14-citropin 1.1.
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| 33369262 |
Rational Design of Helix-Stabilized Antimicrobial Peptide Foldamers Containing α,α-Disubstituted Amino Acids or Side-Chain Stapling |
10.1002/cplu.202000749. |
Chempluschem |
Rational Design of Helix-Stabilized Antimicrobial Peptide Foldamers Containing α,α-Disubstituted Amino Acids or Side-Chain Stapling
Abstract
- Antimicrobial peptides (AMPs) are expected to be good candidate molecules for novel antimicrobial therapies. Most AMPs exert their antimicrobial activity through disruption of microbial membranes due to their amphipathic properties. Recently, the helical peptide 'Stripe' was reported by our group, a rationally designed amphipathic AMP focused on distribution of natural cationic and hydrophobic amino acid residues. In this study, a set of Stripe-based AMP foldamers was designed, synthesized and investigated that contain α,α-disubstituted amino acids or side-chain stapling to stabilize their helical structures. Our results showed that a peptide containing 2-aminoisobutyric acid (Aib) residues exhibited potent antimicrobial activity against both Gram-positive S.aureus (MIC value: 3.125 μM) and Gram-negative bacteria (including a multidrug-resistant strain, MDRP, MIC value: 1.56 μM), without significant hemolytic activity (>100 μM). Electrophysiological measurements revealed that this peptide formed stable pores in a 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine (DOPE)/1,2-dioleoyl-sn-glycero-3-phosphoglycerol (DOPG) bilayer but not in a dioleoylphosphocholine (DOPC) bilayer. The introduction of Aib residues into Stripe could be a promising way to increase the antimicrobial activity of AMP foldamers, and the peptide could represent a promising novel therapeutic candidate to treat multidrug-resistant bacterial infection.
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| 33374547 |
Repurposing the McoTI-II Rigid Molecular Scaffold in to Inhibitor of 'Papain Superfamily' Cysteine Proteases |
10.3390/ph14010007. |
Pharmaceuticals (Basel) |
Repurposing the McoTI-II Rigid Molecular Scaffold in to Inhibitor of 'Papain Superfamily' Cysteine Proteases
Abstract
- Clan C1A or 'papain superfamily' cysteine proteases are key players in many important physiological processes and diseases in most living systems. Novel approaches towards the development of their inhibitors can open new avenues in translational medicine. Here, we report a novel design of a re-engineered chimera inhibitor Mco-cysteine protease inhibitor (CPI) to inhibit the activity of C1A cysteine proteases. This was accomplished by grafting the cystatin first hairpin loop conserved motif (QVVAG) onto loop 1 of the ultrastable cyclic peptide scaffold McoTI-II. The recombinantly expressed Mco-CPI protein was able to bind with micromolar affinity to papain and showed remarkable thermostability owing to the formation of multi-disulphide bonds. Using an in silico approach based on homology modelling, protein-protein docking, the calculation of the free-energy of binding, the mechanism of inhibition of Mco-CPI against representative C1A cysteine proteases (papain and cathepsin L) was validated. Furthermore, molecular dynamics simulation of the Mco-CPI-papain complex validated the interaction as stable. To conclude, in this McoTI-II analogue, the specificity had been successfully redirected towards C1A cysteine proteases while retaining the moderate affinity. The outcomes of this study pave the way for further modifications of the Mco-CPI design for realizing its full potential in therapeutics. This study also demonstrates the relevance of ultrastable peptide-based scaffolds for the development of novel inhibitors via grafting.
|
| 33397096 |
Cyclotides from Brazilian and Their Effects on Human Lymphocytes |
10.1021/acs.jnatprod.0c01069. |
J Nat Prod |
Cyclotides from Brazilian and Their Effects on Human Lymphocytes
Abstract
- Cyclotides are plant-derived peptides found within five families of flowering plants (Violaceae, Rubiaceae, Fabaceae, Solanaceae, and Poaceae) that have a cyclic backbone and six conserved cysteine residues linked by disulfide bonds. Their presence within the Violaceae species seems ubiquitous, yet not all members of other families produce these macrocyclic peptides. The genus Aubl. (Rubiaceae) contains hundreds of neotropical species of shrubs and small trees; however, only a few cyclotides have been discovered hitherto. Herein, five previously uncharacterized Möbius cyclotides within and their pharmacological activities are described. Cyclotides were isolated from leaves and stems of this plant and identified as pase AE, as well as the known peptide kalata S. Cyclotides were de novo sequenced by MALDI-TOF/TOF mass spectrometry, and their structures were solved by NMR spectroscopy. Because some cyclotides have been reported to modulate immune cells, pase AD were assayed for cell proliferation of human primary activated T lymphocytes, and the results showed a dose-dependent antiproliferative function. The toxicity on other nonimmune cells was also assessed. This study reveals that pase cyclotides have potential for applications as immunosuppressants and in immune-related disorders.
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| 33466998 |
Development of Antimicrobial Stapled Peptides Based on Magainin 2 Sequence |
10.3390/molecules26020444. |
Molecules |
Development of Antimicrobial Stapled Peptides Based on Magainin 2 Sequence
Abstract
- Magainin 2 (), which was isolated from the skin of the African clawed frog, is a representative antimicrobial peptide (AMP) that exerts antimicrobial activity via microbial membrane disruption. It has been reported that the helicity and amphipathicity of play important roles in its antimicrobial activity. We investigated and recently reported that 17 amino acid residues of are required for its antimicrobial activity, and accordingly developed antimicrobial foldamers containing α,α-disubstituted amino acid residues. In this study, we further designed and synthesized a set of derivatives bearing the hydrocarbon stapling side chain for helix stabilization. The preferred secondary structures, antimicrobial activities, and cell-membrane disruption activities of the synthesized peptides were evaluated. Our analyses revealed that hydrocarbon stapling strongly stabilized the helical structure of the peptides and enhanced their antimicrobial activity. Moreover, peptide stapling between the first and fifth position from the -terminus showed higher antimicrobial activity than that of against both gram-positive and gram-negative bacteria without exerting significant hemolytic activity. To investigate the modes of action of tested peptides and in antimicrobial and hemolytic activity, electrophysiological measurements were performed.
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| 33570395 |
In Planta Discovery and Chemical Synthesis of Bracelet Cystine Knot Peptides from |
10.1021/acs.jnatprod.0c01065. |
J Nat Prod |
In Planta Discovery and Chemical Synthesis of Bracelet Cystine Knot Peptides from
Abstract
- Cyclotides are plant-derived peptides that have attracted interest as biocides and scaffolds for the development of stable peptide therapeutics. Cyclotides are characterized by their cyclic backbone and cystine knot framework, which engenders them with remarkably high stability. This study reports the cystine knot-related peptidome of , a small rainforest tree in the Violaceae family that is distributed from Australia westward to India. Surprisingly, many more acyclic knotted peptides (acyclotides) were discovered than cyclic counterparts (cyclotides), with 32 acyclotides and 1 cyclotide sequenced using combined transcriptome and proteomic analyses. Nine acyclotides were isolated and screened against a panel of mammalian cell lines, showing they had the cytotoxic properties normally associated with cyclotide-like peptides. NMR analysis of the acyclotide ribes 21 and 22 and the cyclotide ribe 33 confirmed that these peptides contained the cystine knot structural motif. The bracelet-subfamily cyclotide ribe 33 was amenable to chemical synthesis in reasonable yield, an achievement that has long eluded previous attempts to synthetically produce bracelet cyclotides. Accordingly, ribe 33 represents an exciting new bracelet cyclotide scaffold that can be subject to chemical modification for future molecular engineering applications.
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