ABSTRACT
Two complementary approaches were used in search of the intracellular targets of the toxic PR poly-dipeptide encoded by the repeat sequences expanded in the C9orf72 form of amyotrophic lateral sclerosis. The top categories of PRn-bound proteins include constituents of non-membrane invested cellular organelles and intermediate filaments. PRn targets are enriched for the inclusion of low complexity (LC) sequences. Evidence is presented indicating that LC sequences represent the direct target of PRn binding and that interaction between the PRn poly-dipeptide and LC domains is polymer-dependent. These studies indicate that PRn-mediated toxicity may result from broad impediments to the dynamics of cell structure and information flow from gene to message to protein.
Subject(s)
Amyotrophic Lateral Sclerosis/metabolism , Dipeptides/metabolism , Frontotemporal Dementia/metabolism , Peptides/metabolism , Proteins/metabolism , Amyotrophic Lateral Sclerosis/genetics , C9orf72 Protein , DNA Repeat Expansion , Dipeptides/chemistry , Dipeptides/genetics , Frontotemporal Dementia/genetics , HeLa Cells , Humans , Intermediate Filament Proteins/metabolism , Peptides/chemistry , Peptides/genetics , Protein Domains , Proteins/geneticsABSTRACT
HIV can infect non-dividing cells because the viral capsid can overcome the selective barrier of the nuclear pore complex and deliver the genome directly into the nucleus1,2. Remarkably, the intact HIV capsid is more than 1,000 times larger than the size limit prescribed by the diffusion barrier of the nuclear pore3. This barrier in the central channel of the nuclear pore is composed of intrinsically disordered nucleoporin domains enriched in phenylalanine-glycine (FG) dipeptides. Through multivalent FG interactions, cellular karyopherins and their bound cargoes solubilize in this phase to drive nucleocytoplasmic transport4. By performing an in vitro dissection of the nuclear pore complex, we show that a pocket on the surface of the HIV capsid similarly interacts with FG motifs from multiple nucleoporins and that this interaction licences capsids to penetrate FG-nucleoporin condensates. This karyopherin mimicry model addresses a key conceptual challenge for the role of the HIV capsid in nuclear entry and offers an explanation as to how an exogenous entity much larger than any known cellular cargo may be able to non-destructively breach the nuclear envelope.
Subject(s)
Capsid Proteins , Glycine , HIV , Karyopherins , Molecular Mimicry , Nuclear Pore Complex Proteins , Nuclear Pore , Phenylalanine , Humans , Active Transport, Cell Nucleus , Capsid Proteins/chemistry , Capsid Proteins/metabolism , Diffusion , Dipeptides/chemistry , Dipeptides/metabolism , Glycine/metabolism , HIV/chemistry , HIV/metabolism , In Vitro Techniques , Intrinsically Disordered Proteins/chemistry , Intrinsically Disordered Proteins/metabolism , Karyopherins/metabolism , Nuclear Pore/chemistry , Nuclear Pore/metabolism , Nuclear Pore/virology , Nuclear Pore Complex Proteins/chemistry , Nuclear Pore Complex Proteins/metabolism , Permeability , Phenylalanine/metabolism , Solubility , Virus Internalization , Capsid/chemistry , Capsid/metabolismABSTRACT
Hexanucleotide repeat expansion in C9orf72 is one of the most common causes of amyotrophic lateral sclerosis and frontotemporal dementia. The hexanucleotide expansion, formed by GGGGCC (G4C2) repeats, leads to the production of five dipeptide protein repeats (DPRs) via repeat-associated non-AUG translation. Among the five dipeptide repeats, Gly-Arg, Pro-Arg, and Gly-Ala form neuronal inclusions that contain aggregates of the peptides. Several studies have attempted to model DPR-associated toxicity using various repeat lengths, which suggests a unique conformation that is cytotoxic and is independent of the repeat length. However, the structural characteristics of DPR aggregates have yet to be determined. Increasing evidence suggests that soluble species, such as oligomers, are the main cause of toxicity in proteinopathies, such as Alzheimer's and Parkinson's disease. To investigate the ability of DPRs to aggregate and form toxic oligomers, we adopted a reductionist approach using small dipeptide repeats of 3, 6, and 12. This study shows that DPRs, particularly glycine-arginine and proline-arginine, form oligomers that exhibit distinct dye-binding properties and morphologies. Importantly, we also identified toxic DPR oligomers in amyotrophic lateral sclerosis and frontotemporal dementia postmortem brains that are morphologically similar to those generated recombinantly. This study demonstrates that, similar to soluble oligomers formed by various amyloid proteins, DPR oligomers are toxic, independent of their repeat length.
Subject(s)
Amyotrophic Lateral Sclerosis , Frontotemporal Dementia , Humans , Frontotemporal Dementia/genetics , Amyotrophic Lateral Sclerosis/genetics , Amyotrophic Lateral Sclerosis/metabolism , C9orf72 Protein/genetics , C9orf72 Protein/metabolism , DNA Repeat Expansion , Dipeptides/chemistry , Arginine , Amyloidogenic Proteins , GlycineABSTRACT
How organisms integrate metabolism with the external environment is a central question in biology. Here, we describe a novel regulatory small molecule, a proteogenic dipeptide Tyr-Asp, which improves plant tolerance to oxidative stress by directly interfering with glucose metabolism. Specifically, Tyr-Asp inhibits the activity of a key glycolytic enzyme, glyceraldehyde 3-phosphate dehydrogenase (GAPC), and redirects glucose toward pentose phosphate pathway (PPP) and NADPH production. In line with the metabolic data, Tyr-Asp supplementation improved the growth performance of both Arabidopsis and tobacco seedlings subjected to oxidative stress conditions. Moreover, inhibition of Arabidopsis phosphoenolpyruvate carboxykinase (PEPCK) activity by a group of branched-chain amino acid-containing dipeptides, but not by Tyr-Asp, points to a multisite regulation of glycolytic/gluconeogenic pathway by dipeptides. In summary, our results open the intriguing possibility that proteogenic dipeptides act as evolutionarily conserved small-molecule regulators at the nexus of stress, protein degradation, and metabolism.
Subject(s)
Arabidopsis/drug effects , Dipeptides/pharmacology , Glyceraldehyde-3-Phosphate Dehydrogenases/antagonists & inhibitors , Nicotiana/drug effects , Plant Proteins/metabolism , Arabidopsis/metabolism , Arabidopsis Proteins/chemistry , Arabidopsis Proteins/metabolism , Computer Simulation , Dipeptides/chemistry , Dipeptides/metabolism , Glyceraldehyde-3-Phosphate Dehydrogenase (Phosphorylating)/chemistry , Glyceraldehyde-3-Phosphate Dehydrogenase (Phosphorylating)/metabolism , Glyceraldehyde-3-Phosphate Dehydrogenases/metabolism , NADP/metabolism , Oxidation-Reduction , Oxidative Stress/drug effects , Pentose Phosphate Pathway/drug effects , Phosphoenolpyruvate Carboxykinase (ATP)/metabolism , Plant Proteins/antagonists & inhibitors , Seedlings/drug effects , Seedlings/metabolism , Nicotiana/metabolismABSTRACT
Amide bond formation is one of the most important reactions in both chemistry and biology1-4, but there is currently no chemical method of achieving α-peptide ligation in water that tolerates all of the 20 proteinogenic amino acids at the peptide ligation site. The universal genetic code establishes that the biological role of peptides predates life's last universal common ancestor and that peptides played an essential part in the origins of life5-9. The essential role of sulfur in the citric acid cycle, non-ribosomal peptide synthesis and polyketide biosynthesis point towards thioester-dependent peptide ligations preceding RNA-dependent protein synthesis during the evolution of life5,9-13. However, a robust mechanism for aminoacyl thioester formation has not been demonstrated13. Here we report a chemoselective, high-yielding α-aminonitrile ligation that exploits only prebiotically plausible molecules-hydrogen sulfide, thioacetate12,14 and ferricyanide12,14-17 or cyanoacetylene8,14-to yield α-peptides in water. The ligation is extremely selective for α-aminonitrile coupling and tolerates all of the 20 proteinogenic amino acid residues. Two essential features enable peptide ligation in water: the reactivity and pKaH of α-aminonitriles makes them compatible with ligation at neutral pH and N-acylation stabilizes the peptide product and activates the peptide precursor to (biomimetic) N-to-C peptide ligation. Our model unites prebiotic aminonitrile synthesis and biological α-peptides, suggesting that short N-acyl peptide nitriles were plausible substrates during early evolution.
Subject(s)
Evolution, Chemical , Nitriles/chemistry , Nitriles/chemical synthesis , Origin of Life , Peptides/chemistry , Peptides/chemical synthesis , Water/chemistry , Acetylene/analogs & derivatives , Acetylene/chemistry , Dipeptides/chemical synthesis , Dipeptides/chemistry , Ferricyanides/chemistry , Hydrogen Sulfide/chemistry , Hydrogen-Ion Concentration , Oxidation-Reduction , Sulfhydryl Compounds/chemistry , Sulfides/chemistryABSTRACT
Liquid-liquid phase separation (LLPS) of RNA-binding proteins plays an important role in the formation of multiple membrane-less organelles involved in RNA metabolism, including stress granules. Defects in stress granule homeostasis constitute a cornerstone of ALS/FTLD pathogenesis. Polar residues (tyrosine and glutamine) have been previously demonstrated to be critical for phase separation of ALS-linked stress granule proteins. We now identify an active role for arginine-rich domains in these phase separations. Moreover, arginine-rich dipeptide repeats (DPRs) derived from C9orf72 hexanucleotide repeat expansions similarly undergo LLPS and induce phase separation of a large set of proteins involved in RNA and stress granule metabolism. Expression of arginine-rich DPRs in cells induced spontaneous stress granule assembly that required both eIF2α phosphorylation and G3BP. Together with recent reports showing that DPRs affect nucleocytoplasmic transport, our results point to an important role for arginine-rich DPRs in the pathogenesis of C9orf72 ALS/FTLD.
Subject(s)
Amyotrophic Lateral Sclerosis/metabolism , Arginine/metabolism , Cytoplasmic Granules/metabolism , Dipeptides/metabolism , Intrinsically Disordered Proteins/metabolism , Proteins/metabolism , Amyotrophic Lateral Sclerosis/genetics , Amyotrophic Lateral Sclerosis/pathology , Arginine/chemistry , C9orf72 Protein , Carrier Proteins/genetics , Carrier Proteins/metabolism , Cytoplasmic Granules/pathology , DNA Helicases , Dipeptides/chemistry , Eukaryotic Initiation Factor-2/genetics , Eukaryotic Initiation Factor-2/metabolism , HeLa Cells , Humans , Intrinsically Disordered Proteins/chemistry , Lipid Droplets/metabolism , Phosphorylation , Poly-ADP-Ribose Binding Proteins , Protein Domains , Proteins/chemistry , RNA/metabolism , RNA Helicases , RNA Recognition Motif Proteins , Time Factors , TransfectionABSTRACT
Amide bond formation, the essential condensation reaction underlying peptide synthesis, is hindered in aqueous systems by the thermodynamic constraints associated with dehydration. This represents a key difficulty for the widely held view that prebiotic chemical evolution leading to the formation of the first biomolecules occurred in an oceanic environment. Recent evidence for the acceleration of chemical reactions at droplet interfaces led us to explore aqueous amino acid droplet chemistry. We report the formation of dipeptide isomer ions from free glycine or L-alanine at the air-water interface of aqueous microdroplets emanating from a single spray source (with or without applied potential) during their flight toward the inlet of a mass spectrometer. The proposed isomeric dipeptide ion is an oxazolidinone that takes fully covalent and ion-neutral complex forms. This structure is consistent with observed fragmentation patterns and its conversion to authentic dipeptide ions upon gentle collisions and for its formation from authentic dipeptides at ultra-low concentrations. It also rationalizes the results of droplet fusion experiments that show that the dipeptide isomer facilitates additional amide bond formation events, yielding authentic tri- through hexapeptides. We propose that the interface of aqueous microdroplets serves as a drying surface that shifts the equilibrium between free amino acids in favor of dehydration via stabilization of the dipeptide isomers. These findings offer a possible solution to the water paradox of biopolymer synthesis in prebiotic chemistry.
Subject(s)
Amino Acids , Oxazolidinones , Alanine , Amides , Amino Acids/chemistry , Biopolymers , Dehydration , Dipeptides/chemistry , Glycine , Humans , Peptides/chemistry , Water/chemistryABSTRACT
Hexanucleotide expansion mutations in C9ORF72 are a frequent cause of amyotrophic lateral sclerosis. We previously reported that long arginine-rich dipeptide repeats (DPRs), mimicking abnormal proteins expressed from the hexanucleotide expansion, caused translation stalling when expressed in cell culture models. Whether this stalling provides a mechanism of pathogenicity remains to be determined. Here, we explored the molecular features of DPR-induced stalling and examined whether known mechanisms such as ribosome quality control (RQC) regulate translation elongation on sequences that encode arginine-rich DPRs. We demonstrate that arginine-rich DPRs lead to stalling in a length-dependent manner, with lengths longer than 40 repeats invoking severe translation arrest. Mutational screening of 40×Gly-Xxx DPRs shows that stalling is most pronounced when Xxx is a charged amino acid (Arg, Lys, Glu, or Asp). Through a genome-wide knockout screen, we find that genes regulating stalling on polyadenosine mRNA coding for poly-Lys, a canonical RQC substrate, act differently in the case of arginine-rich DPRs. Indeed, these findings point to a limited scope for natural regulatory responses to resolve the arginine-rich DPR stalls, even though the stalls may be sensed, as evidenced by an upregulation of RQC gene expression. These findings therefore implicate arginine-rich DPR-mediated stalled ribosomes as a source of stress and toxicity and may be a crucial component in pathomechanisms.
Subject(s)
Amyotrophic Lateral Sclerosis , Humans , Amyotrophic Lateral Sclerosis/genetics , Amyotrophic Lateral Sclerosis/physiopathology , Arginine/metabolism , C9orf72 Protein/genetics , C9orf72 Protein/metabolism , Dipeptides/chemistry , Ribosomes/genetics , Ribosomes/metabolism , Gene Knockout Techniques , Mutation , Up-RegulationABSTRACT
Allostery, as seen in extant biology, governs the activity regulation of enzymes through the redistribution of conformational equilibria upon binding an effector. Herein, a minimal design is demonstrated where a dipeptide can exploit dynamic imine linkage to condense with simple aldehydes to access spherical aggregates as catalytically active states, which facilitates an orthogonal reaction due to the closer proximity of catalytic residues (imidazoles). The allosteric site (amine) of the minimal catalyst can concomitantly bind to an inhibitor via a dynamic exchange, which leads to the alternation of the energy landscape of the self-assembled state, resulting in downregulation of catalytic activity. Further, temporal control over allosteric regulation is realized via a feedback-controlled autonomous reaction network that utilizes the hydrolytic activity of the (in)active state as a function of time.
Subject(s)
Dipeptides , Allosteric Regulation , Dipeptides/chemistry , Catalysis , Molecular Structure , BiocatalysisABSTRACT
In an effort to target polypeptides at nonterminal sites, we screened the binding of the synthetic receptor cucurbit[8]uril (Q8) to a small library of tetrapeptides, each containing a nonterminal dipeptide binding site. The resulting leads were characterized in detail using a combination of isothermal titration calorimetry, 1H NMR spectroscopy, electrospray ionization time-of-flight mass spectrometry (ESI-TOF-MS), and X-ray crystallography. The equilibrium dissociation constant values determined for the binding of Q8 to nonterminal dipeptide sites Lys-Phe (KF) and Phe-Lys (FK) were 60 and 86 nm, respectively. These are to the best of our knowledge the highest affinities reported to date for any synthetic receptor targeting a nonterminal site on an unmodified peptide. A 0.79 Å resolution crystal structure was obtained for the complex of Q8 with the peptide Gly-Gly-Leu-Tyr-Gly-Gly-Gly (GGLYGGG) and reveals structural details of the pair-inclusion motif. The molecular basis for recognition is established to be the inclusion of the side chains of Leu and Tyr residues, as well as an extensive network of hydrogen bonds between the peptide backbone, the carbonyl oxygens of Q8, and proximal water molecules. In addition, the crystal structure reveals that Q8 induces a type II ß-turn. The sequence-selectivity, high affinity, reversibility, and detailed structural characterization of this system should facilitate the development of applications involving ligand-induced polypeptide folding.
Subject(s)
Receptors, Artificial , Dipeptides/chemistry , Peptides/chemistry , Crystallography, X-Ray , Binding SitesABSTRACT
The development of lipid nanoparticles (LNPs) has enabled the clinical application of small interfering ribonucleic acid (siRNA)-based therapies. Accordingly, various unique ionizable lipids have been explored for efficient siRNA delivery. However, safety concerns related to the structure of ionizable lipids have been raised. Here, we developed a pH-responsive dipeptide-conjugated lipid (DPL) for efficient, high safety siRNA delivery. We synthesized a DPL library by varying the dipeptide sequence and established a strong correlation between the knockdown efficiency of the DPL-based LNPs and the dipeptide sequence. The LNPs prepared with a DPL containing arginine (R) and glutamic acid (E) (DPL-ER) exhibited the highest knockdown efficiency. In addition, the DPL-ER-based LNPs with relatively long lipid tails (DPL-ER-C22:C22) exhibited a higher knockdown efficiency than those with short ones (DPL-ER-18:C18). The zeta potential of the DPL-ER-C22:C22-based LNPs increased as the pH decreased from 7.4 (physiological condition) to 5.5 (endosomal condition). Importantly, the DPL-ER-C22:C22-based LNPs exhibited a higher knockdown efficiency than the LNPs prepared using commercially available ionizable lipids. These results suggest that the DPL-based LNPs are safe and efficient siRNA delivery carriers.
Subject(s)
Dipeptides , Lipids , Nanoparticles , RNA, Small Interfering , Hydrogen-Ion Concentration , RNA, Small Interfering/chemistry , RNA, Small Interfering/administration & dosage , Nanoparticles/chemistry , Lipids/chemistry , Dipeptides/chemistry , Humans , HeLa CellsABSTRACT
The utilization of the glycated amino acids formyline and pyrraline as well as their peptide-bound derivatives by 14 Saccharomyces yeasts, including 6 beer yeasts (bottom and top fermenting), one wine yeast, 6â strains isolated from natural habitats and one laboratory reference yeast strain (wild type) was investigated. All yeasts were able to metabolize glycated amino acids via the Ehrlich pathway to the corresponding Ehrlich metabolites. While formyline and small amounts of pyrraline entered the yeast cells via passive diffusion, the amounts of dipeptide-bound MRPs, especially the dipeptides glycated at the C-terminus, decreased much faster, indicating an uptake into the yeast cells. Furthermore, the glycation-mediated hydrophobization in general leads to an faster degradation rate compared to the native lysine dipeptides. While the utilization of free formyline is yeast-specific, the amounts of (glycated) dipeptides decreased faster in the presence of brewer's yeasts, which also showed a higher formation rate of Ehrlich metabolites compared to naturally isolated strains. Due to rapid uptake of alanyl dipeptides, it can be assumed that the Ehrlich enzyme system of naturally isolated yeasts is overloaded and the intracellularly released MRP is primarily excreted from the cell. This indicates adaptation of technologically used yeasts to (glycated) dipeptides as a nitrogen source.
Subject(s)
Dipeptides , Norleucine , Dipeptides/metabolism , Dipeptides/chemistry , Norleucine/metabolism , Norleucine/analogs & derivatives , Norleucine/chemistry , Saccharomyces/metabolism , Saccharomyces cerevisiae/metabolism , Glycosylation , PyrrolesABSTRACT
Dipeptides of a new structure based on ß-triazolalanines and (L)-α-amino acids were synthesized and optimal conditions were developed that ensure both chemical and optical purity of the final products. Molecular docking was carried out and possible intermolecular interactions of dipeptides with potential targets were established. Based on these studies, the analgesic property of chosen dipeptides was studied and it was found that some compounds possess revealed antinociceptive activity in the tail-flick test.
Subject(s)
Analgesics , Dipeptides , Molecular Docking Simulation , Triazoles , Analgesics/chemistry , Analgesics/pharmacology , Analgesics/chemical synthesis , Triazoles/chemistry , Triazoles/pharmacology , Triazoles/chemical synthesis , Dipeptides/chemistry , Dipeptides/chemical synthesis , Dipeptides/pharmacology , Animals , Mice , MaleABSTRACT
The synthesis of linker-payloads is a critical step in developing antibody-drug conjugates (ADCs), a rapidly advancing therapeutic approach in oncology. The conventional method for synthesizing cathepsin B-labile dipeptide linkers, which are commonly used in ADC development, involves the solution-phase assembly of cathepsin B-sensitive dipeptides, followed by the installation of self-immolative para-aminobenzyl carbonate to facilitate the attachment of potent cytotoxic payloads. However, this approach is often low yield and laborious, especially when extending the peptide chain with components like glutamic acid to improve mouse serum stability or charged amino acids or poly(ethylene glycol) moieties to enhance linker hydrophilicity. Here, we introduce a novel approach utilizing late-stage desulfurization chemistry, enabling safe, facile, and cost-effective access to the cathepsin B-cleavable linker, Val-Ala-PABC-MMAE, on resin for the first time.
Subject(s)
Cathepsin B , Immunoconjugates , Oligopeptides , Solid-Phase Synthesis Techniques , Immunoconjugates/chemistry , Cathepsin B/metabolism , Solid-Phase Synthesis Techniques/methods , Oligopeptides/chemistry , Humans , Animals , Mice , Dipeptides/chemistryABSTRACT
PURPOSE: A series of new 68Ga-labeled tracers based on [68Ga]Ga-PSMA-617 were developed to augment the tumor-to-kidney ratio and reduce the activity accumulation in bladder, ultimately minimize radiation toxicity to the urinary system. METHODS: We introduced quinoline group, phenylalanine and decanoic acid into different tracers to enhance their lipophilicity, strategically limiting their metabolic pathway through the urinary system. Their binding affinity onto LNCaP cells was determined through in vitro saturation assays and competition binding assays. In vivo metabolic study, PET imaging and biodistribution experiment were performed in LNCaP tumor-bearing B-NSG male mice. The most promising tracer was selected for first-in-human study. RESULTS: Four radiotracers were synthesized with radiochemical purity (RCP) > 95% and molar activity in a range of 20.0-25.5 GBq/µmol. The binding affinities (Ki) of TWS01, TWS02 to PSMA were in the low nanomolar range (< 10 nM), while TWS03 and TWS04 exhibited binding affinities with Ki > 20 nM (59.42 nM for TWS03 and 37.14 nM for TWS04). All radiotracers exhibited high stability in vivo except [68Ga]Ga-TWS03. Micro PET/CT imaging and biodistribution analysis revealed that [68Ga]Ga-TWS02 enabled clear tumor visualization in PET images at 1.5 h post-injection, with higher tumor-to-kidney ratio (T/K, 0.93) and tumor-to-muscle ratio (T/M, 107.62) compared with [68Ga]Ga-PSMA-617 (T/K: 0.39, T/M: 15.01) and [68Ga]Ga-PSMA-11 (T/K: 0.15, T/M: 24.00). In first-in-human study, [68Ga]Ga-TWS02 effectively detected PCa-associated lesions including primary and metastatic lesions, with lower accumulation in urinary system, suggesting that [68Ga]Ga-TWS02 might be applied in the detection of bladder invasion, with minimized radiation toxicity to the urinary system. CONCLUSION: Introduction of quinoline group, phenylalanine and decanoic acid into different tracers can modulate the binding affinity and pharmacokinetics of PSMA in vivo. [68Ga]Ga-TWS02 showed high binding affinity to PSMA, excellent pharmacokinetic properties and clear imaging of PCa-associated lesions, making it a promising radiotracer for the clinical diagnosis of PCa. Moreover, TWS02 with a chelator DOTA could also label 177Lu and 225Ac, which could be used for PCa treatment without significant side effects. TRIAL REGISTRATION: The clinical evaluation of this study was registered On October 30, 2021 at https://www.chictr.org.cn/ (No: ChiCTR2100052545).
Subject(s)
Glutamate Carboxypeptidase II , Positron-Emission Tomography , Humans , Male , Mice , Animals , Tissue Distribution , Cell Line, Tumor , Glutamate Carboxypeptidase II/metabolism , Positron-Emission Tomography/methods , Radioactive Tracers , Gallium Radioisotopes/pharmacokinetics , Prostatic Neoplasms/diagnostic imaging , Prostatic Neoplasms/metabolism , Prostatic Neoplasms/radiotherapy , Antigens, Surface/metabolism , Radiopharmaceuticals/pharmacokinetics , Radiopharmaceuticals/chemistry , Radiochemistry , Dipeptides/pharmacokinetics , Dipeptides/chemistry , Heterocyclic Compounds, 1-Ring/chemistry , Heterocyclic Compounds, 1-Ring/pharmacokinetics , Positron Emission Tomography Computed Tomography/methodsABSTRACT
The abiotic synthesis of peptides, widely regarded as one of the key chemical reactions on the prebiotic Earth, is thermodynamically constrained in solution. Herein, a simulation of the lightning phenomenon on the sea surface using bubble bursting and arc plasma under ambient conditions enables dipeptide formation of six amino acids with conversion ratios ranging from 2.6 % to 25.5 %. Additionally, we observed the formation of biologically active tripeptides and investigated the stereoselectivity of the dipeptide formation reaction. By utilizing a mixture of 20 amino acids in the reaction, 102 possible dipeptides were generated. These results establish experimental constructions to mimic achievable prebiotic conditions and provide a credible pathway for endogenous biopolymer synthesis on prebiotic Earth.
Subject(s)
Dipeptides , Peptides , Peptides/chemistry , Dipeptides/chemistry , Amino Acids/chemistry , Thermodynamics , Stereoisomerism , Plasma Gases/chemistry , Prebiotics , Origin of LifeABSTRACT
Cyclic dipeptides (CDPs) are crucial building blocks for a range of functional nanomaterials due to their simple chemical structure and high molecular stability. In this investigation, we synthesized a set of S-benzyl-L-cysteine-based CDPs (designated as P1-P6) and thoroughly examined their self-assembly behavior in a methanol-water solvent to elucidate the relationship between their structure and gelation properties. The hydrophobicity of the amino acids within the CDPs was gradually increased. The present study employed a comprehensive array of analytical techniques, including NMR, FT-IR, AFM, thioflavin-T, congo-red CD, X-ray crystallography, and biophysical calculations like Hirshfield Surface analysis and DFT analysis. These methods revealed that in addition to hydrogen bonding, the hydrophobic nature of the amino acid side chain significantly influences the propensity of CDPs to form hydrogels. Each CDP yielded distinct nanofibrillar networks rich in ß-sheet structures, showcasing unique morphological features. Moreover, we explored the practical application of these CDP-based hydrogels in water purification by utilizing them to remove harmful organic dyes from contaminated water. This application underscores the potential of CDPs in addressing environmental challenges, offering a promising avenue for the future development of these materials in water treatment technologies.
Subject(s)
Cysteine , Dipeptides , Hydrogels , Nanostructures , Peptides, Cyclic , Dipeptides/chemistry , Cysteine/chemistry , Hydrogels/chemistry , Peptides, Cyclic/chemistry , Nanostructures/chemistry , Crystallography, X-Ray , Hydrophobic and Hydrophilic Interactions , Hydrogen Bonding , Water Purification/methods , Spectroscopy, Fourier Transform InfraredABSTRACT
Transactive response DNA-binding Protein of 43 kDa (TDP-43) assembles various aggregate forms, including biomolecular condensates or functional and pathological amyloids, with roles in disparate scenarios (e.g., muscle regeneration versus neurodegeneration). The link between condensates and fibrils remains unclear, just as the factors controlling conformational transitions within these aggregate species: Salt- or RNA-induced droplets may evolve into fibrils or remain in the droplet form, suggesting distinct end point species of different aggregation pathways. Using microscopy and NMR methods, we unexpectedly observed in vitro droplet formation in the absence of salts or RNAs and provided visual evidence for fibrillization at the droplet surface/solvent interface but not the droplet interior. Our NMR analyses unambiguously uncovered a distinct amyloid conformation in which Phe-Gly motifs are key elements of the reconstituted fibril form, suggesting a pivotal role for these residues in creating the fibril core. This contrasts the minor participation of Phe-Gly motifs in initiation of the droplet form. Our results point to an intrinsic (i.e., non-induced) aggregation pathway that may exist over a broad range of conditions and illustrate structural features that distinguishes between aggregate forms.
Subject(s)
DNA-Binding Proteins/chemistry , DNA-Binding Proteins/metabolism , Dipeptides/chemistry , Protein Aggregates , Amino Acid Sequence , Amyloid/chemistry , Amyloid/metabolism , Chemical Precipitation , Dipeptides/physiology , Humans , Hydrogen-Ion Concentration , Protein Aggregation, Pathological/metabolism , Protein Aggregation, Pathological/pathology , Protein Interaction Domains and Motifs/physiology , Solvents/chemistry , Solvents/pharmacologyABSTRACT
Nitric oxide (NO) donating drugs such as organic nitrates have been used to treat cardiovascular diseases for more than a century. These donors primarily produce NO systemically. It is however sometimes desirable to control the amount, location, and time of NO delivery. We present the design of a novel pH-sensitive NO release system that is achieved by the synthesis of dipeptide diphenylalanine (FF) and graphene oxide (GO) co-assembled hybrid nanosheets (termed as FF@GO) through weak molecular interactions. These hybrid nanosheets were characterised by using X-ray diffraction, Raman spectroscopy, Fourier transform infrared spectroscopy, zeta potential measurements, X-ray photoelectron spectroscopy, scanning and transmission electron microscopies. The weak molecular interactions, which include electrostatic, hydrogen bonding and π-π stacking, are pH sensitive due to the presence of carboxylic acid and amine functionalities on GO and the dipeptide building blocks. Herein, we demonstrate that this formulation can be loaded with NO gas with the dipeptide acting as an arresting agent to inhibit NO burst release at neutral pH; however, at acidic pH it is capable of releasing NO at the rate of up to 0.6 µM per minute, comparable to the amount of NO produced by healthy endothelium. In conclusion, the innovative conjugation of dipeptide with graphene can store and release NO gas under physiologically relevant concentrations in a pH-responsive manner. pH responsive NO-releasing organic-inorganic nanohybrids may prove useful for the treatment of cardiovascular diseases and other pathologies.
Subject(s)
Graphite , Nanostructures , Nitric Oxide , Graphite/chemistry , Hydrogen-Ion Concentration , Nitric Oxide/chemistry , Nitric Oxide/metabolism , Nanostructures/chemistry , Humans , Dipeptides/chemistry , Phenylalanine/chemistry , Phenylalanine/analogs & derivativesABSTRACT
Cells require oligonucleotides and polypeptides with specific, homochiral sequences to perform essential functions, but it is unclear how such oligomers were selected from random sequences at the origin of life. Cells were probably preceded by simple compartments such as fatty acid vesicles, and oligomers that increased the stability, growth, or division of vesicles could have thereby increased in frequency. We therefore tested whether prebiotic peptides alter the stability or growth of vesicles composed of a prebiotic fatty acid. We find that three of 15 dipeptides tested reduce salt-induced flocculation of vesicles. All three contain leucine, and increasing their length increases the efficacy. Also, leucine-leucine but not alanine-alanine increases the size of vesicles grown by multiple additions of micelles. In a molecular simulation, leucine-leucine docks to the membrane, with the side chains inserted into the hydrophobic core of the bilayer, while alanine-alanine fails to dock. Finally, the heterochiral forms of leucine-leucine, at a high concentration, rapidly shrink the vesicles and make them leakier and less stable to high pH than the homochiral forms do. Thus, prebiotic peptide-membrane interactions influence the flocculation, growth, size, leakiness, and pH stability of prebiotic vesicles, with differential effects due to sequence, length, and chirality. These differences could lead to a population of vesicles enriched for peptides with beneficial sequence and chirality, beginning selection for the functional oligomers that underpin life.