RESUMEN

Peripheral artery disease is commonly treated with balloon angioplasty, a procedure involving minimally invasive, transluminal insertion of a catheter to the site of stenosis, where a balloon is inflated to open the blockage, restoring blood flow. However, peripheral angioplasty has a high rate of restenosis, limiting long-term patency. Therefore, angioplasty is sometimes paired with delivery of cytotoxic drugs like paclitaxel to reduce neointimal tissue formation. We pursue intravascular drug delivery strategies that target the underlying cause of restenosis - intimal hyperplasia resulting from stress-induced vascular smooth muscle cell switching from the healthy contractile into a pathological synthetic phenotype. We have established MAPKAP kinase 2 (MK2) as a driver of this phenotype switch and seek to establish convective and contact transfer (coated balloon) methods for MK2 inhibitory peptide delivery to sites of angioplasty. Using a flow loop bioreactor, we showed MK2 inhibition in ex vivo arteries suppresses smooth muscle cell phenotype switching while preserving vessel contractility. A rat carotid artery balloon injury model demonstrated inhibition of intimal hyperplasia following MK2i coated balloon treatment in vivo. These studies establish both convective and drug coated balloon strategies as promising approaches for intravascular delivery of MK2 inhibitory formulations to improve efficacy of balloon angioplasty.

Asunto(s)

Péptidos y Proteínas de Señalización Intracelular , Proteínas Serina-Treonina Quinasas , Ratas Sprague-Dawley , Animales , Proteínas Serina-Treonina Quinasas/metabolismo , Proteínas Serina-Treonina Quinasas/antagonistas & inhibidores , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Masculino , Péptidos/química , Péptidos/farmacología , Ratas , Músculo Liso Vascular/efectos de los fármacos , Músculo Liso Vascular/citología , Angioplastia de Balón/métodos , Miocitos del Músculo Liso/efectos de los fármacos , Miocitos del Músculo Liso/citología , Miocitos del Músculo Liso/metabolismo , Sistemas de Liberación de Medicamentos , Hiperplasia/prevención & control , Angioplastia , Neointima/prevención & control , Neointima/patología

RESUMEN

This study isolated a novel peptide MMGGED with strong calcium-binding capacity from defatted walnut meal and synthesized a novel peptide­calcium chelate COS-MMGGED-Ca with high stability via glycation. Structural characterization and computer simulation identified binding sites, while in vitro digestion stability and calcium transport experiments explored the chelate's properties. Results showed that after glycation, COS-MMGGED bound Ca2+ with 88.75 ± 1.75 %, mainly via aspartic and glutamic acids. COS-MMGGED-Ca released Ca2+ steadily (60.27 %), with thermal denaturation temperature increased by 18 °C and 37 °C compared to MMGGED-Ca, indicating good processing performance. Furthermore, COS-MMGGED significantly enhanced Ca2+ transport across Caco-2 monolayers, 1.13-fold and 1.62-fold higher than CaCl2 and MMGGED, respectively, at 240 h. These findings prove glycation enhances structural properties, stability, calcium loading, and transport of peptide­calcium chelates, providing a scientific basis for developing novel efficient calcium supplements and high-value utilization of walnut meal.

Asunto(s)

Calcio , Juglans , Péptidos , Juglans/química , Humanos , Calcio/química , Calcio/metabolismo , Células CACO-2 , Péptidos/química , Péptidos/metabolismo , Glicosilación , Quelantes del Calcio/química

RESUMEN

Fortification of human milk (HM) is often necessary to meet the nutritional requirements of preterm infants. The present experiment aimed to establish whether the supplementation of HM with either an experimental donkey milk-derived fortifier containing whole donkey milk proteins, or with a commercial bovine milk-derived fortifier containing hydrolyzed bovine whey proteins, affects peptide release differently during digestion. The experiment was conducted using an in vitro dynamic system designed to simulate the preterm infant's digestion followed by digesta analysis by means of LC-MS-MS. The different fortifiers did not appear to influence the cumulative intensity of HM peptides. Fortification had a differential impact on the release of either donkey or bovine bioactive peptides. Donkey milk peptides showed antioxidant/ACE inhibitory activities, while bovine peptides showed opioid, dipeptil- and propyl endo- peptidase inhibitory and antimicrobial activity. A slight delay in peptide release from human lactoferrin and α-lactalbumin was observed when HM was supplemented with donkey milk-derived fortifier.

Asunto(s)

Digestión , Equidae , Proteínas de la Leche , Leche Humana , Péptidos , Humanos , Animales , Leche Humana/química , Leche Humana/metabolismo , Proteínas de la Leche/química , Proteínas de la Leche/metabolismo , Proteínas de la Leche/análisis , Bovinos , Péptidos/química , Péptidos/metabolismo , Alimentos Fortificados/análisis , Espectrometría de Masas en Tándem , Modelos Biológicos , Proteína de Suero de Leche/química , Proteína de Suero de Leche/metabolismo

RESUMEN

The study examined the antihypertensive effect of peptides derived from pepsin-hydrolyzed corn gluten meal, namely KQLLGY and PPYPW, and their in silico gastrointestinal tract digested fragments, KQL and PPY, respectively. KQLLGY and PPYPW showed higher angiotensin I-converting enzyme (ACE)-inhibitory activity and lower ACE inhibition constant (Ki) values when compared to KQL and PPY. Only KQL showed a mild antihypertensive effect in spontaneously hypertensive rats with -7.83 and - 5.71 mmHg systolic and diastolic blood pressure values, respectively, after 8 h oral administration. During passage through Caco-2 cells, KQL was further degraded to QL, which had reduced ACE inhibitory activity. In addition, molecular dynamics revealed that the QL-ACE complex was less stable compared to the KQL-ACE. This study reveals that structural transformation during peptide permeation plays a vital role in attenuating antihypertensive effect of the ACE inhibitor peptide.

Asunto(s)

Inhibidores de la Enzima Convertidora de Angiotensina , Antihipertensivos , Digestión , Glútenes , Péptidos , Peptidil-Dipeptidasa A , Ratas Endogámicas SHR , Zea mays , Inhibidores de la Enzima Convertidora de Angiotensina/química , Inhibidores de la Enzima Convertidora de Angiotensina/farmacología , Inhibidores de la Enzima Convertidora de Angiotensina/metabolismo , Antihipertensivos/química , Antihipertensivos/farmacología , Animales , Glútenes/química , Glútenes/metabolismo , Humanos , Zea mays/química , Zea mays/metabolismo , Ratas , Células CACO-2 , Péptidos/química , Péptidos/farmacología , Masculino , Digestión/efectos de los fármacos , Peptidil-Dipeptidasa A/química , Peptidil-Dipeptidasa A/metabolismo , Presión Sanguínea/efectos de los fármacos , Hipertensión/metabolismo , Hipertensión/tratamiento farmacológico , Hipertensión/fisiopatología , Tracto Gastrointestinal/metabolismo , Hidrolisados de Proteína/química , Hidrolisados de Proteína/farmacología , Hidrólisis

RESUMEN

Mesenchymal stem cells (MSCs) are expected to be useful therapeutics in osteoarthritis (OA), the most common joint disorder characterized by cartilage degradation. However, evidence is limited with regard to cartilage repair in clinical trials because of the uncontrolled differentiation and weak cartilage-targeting ability of MSCs after injection. To overcome these drawbacks, here we synthesized CuO@MSN nanoparticles (NPs) to deliver Sox9 plasmid DNA (favoring chondrogenesis) and recombinant protein Bmp7 (inhibiting hypertrophy). After taking up CuO@MSN/Sox9/Bmp7 (CSB NPs), the expressions of chondrogenic markers were enhanced while hypertrophic markers were decreased in response to these CSB-engineered MSCs. Moreover, a cartilage-targeted peptide (designated as peptide W) was conjugated onto the surface of MSCs via a click chemistry reaction, thereby prolonging the residence time of MSCs in both the knee joint cavity of mice and human-derived cartilage. In a surgery-induced OA mouse model, the NP and peptide dual-modified W-CSB-MSCs showed an enhancing therapeutic effect on cartilage repair in knee joints compared with other engineered MSCs after intra-articular injection. Most importantly, W-CSB-MSCs accelerated cartilage regeneration in damaged cartilage explants derived from OA patients. Thus, this new peptide and NPs dual engineering strategy shows potential for clinical applications to boost cartilage repair in OA using MSC therapy.

Asunto(s)

Diferenciación Celular , Trasplante de Células Madre Mesenquimatosas , Células Madre Mesenquimatosas , Nanopartículas , Osteoartritis , Péptidos , Células Madre Mesenquimatosas/citología , Células Madre Mesenquimatosas/metabolismo , Animales , Osteoartritis/terapia , Osteoartritis/patología , Nanopartículas/química , Humanos , Diferenciación Celular/efectos de los fármacos , Péptidos/química , Trasplante de Células Madre Mesenquimatosas/métodos , Condrogénesis/efectos de los fármacos , Ratones , Factor de Transcripción SOX9/metabolismo , Factor de Transcripción SOX9/genética , Cartílago Articular/patología , Cartílago Articular/efectos de los fármacos , Proteína Morfogenética Ósea 7/química , Proteína Morfogenética Ósea 7/farmacología , Ingeniería de Tejidos/métodos , Regeneración/efectos de los fármacos

RESUMEN

BACKGROUND: Mass spectrometry (MS)-based proteomics is a powerful tool for identifying and quantifying proteins. However, chimeric spectra caused by the fragmentation of multiple precursors within the same isolation window impair the accuracy of peptide identification and isobaric mass tag-based quantification. While there have been advances in computational deconvolution of chimeric spectra and methods to further separate the peptides by ion mobility or through MSn, the use of narrower isolation windows to decrease the fraction of chimeric species remains to be fully explored. RESULTS: We present results obtained on a SCIEX TripleTOF instrument where the quadrupole was optimized and tuned for precursor isolation at 0.1 Da (FWHH). Using a three-proteome model (trypsin digest of protein lysates from yeast, human and E. coli) and 8-plex iTRAQ labeling to document the interference effect, we investigated the impact of co-fragmentation on spectral purity, identification accuracy and quantification accuracy. The narrow quadrupole isolation window significantly improved the spectral purity and reduced the interference of non-target precursors on quantification accuracy. The high-resolution isolation strategy also reduced the number of false identifications caused by chimeric spectra. While these improvements came at the cost of sensitivity loss, combining high-resolution isolation with other advanced techniques, including ion mobility, may result in improved accuracy in identification and quantification. SIGNIFICANCE: Compared to standard-resolution quadrupole isolation (0.7 Da), high-resolution quadrupole isolation (0.1 Da) significantly improved the spectral purity and quantification accuracy while reducing the number of potential false identifications caused by chimeric spectra, thus showing excellent potential for further development to analyze clinical proteomics samples, for which high accuracy is essential.

Asunto(s)

Proteómica , Proteómica/métodos , Humanos , Iones/química , Escherichia coli/química , Saccharomyces cerevisiae/química , Péptidos/química , Péptidos/análisis , Espectrometría de Masas/métodos

RESUMEN

Anticancer peptides (ACPs), naturally occurring molecules with remarkable potential to target and kill cancer cells. However, identifying ACPs based solely from their primary amino acid sequences remains a major hurdle in immunoinformatics. In the past, several web-based machine learning (ML) tools have been proposed to assist researchers in identifying potential ACPs for further testing. Notably, our meta-approach method, mACPpred, introduced in 2019, has significantly advanced the field of ACP research. Given the exponential growth in the number of characterized ACPs, there is now a pressing need to create an updated version of mACPpred. To develop mACPpred 2.0, we constructed an up-to-date benchmarking dataset by integrating all publicly available ACP datasets. We employed a large-scale of feature descriptors, encompassing both conventional feature descriptors and advanced pre-trained natural language processing (NLP)-based embeddings. We evaluated their ability to discriminate between ACPs and non-ACPs using eleven different classifiers. Subsequently, we employed a stacked deep learning (SDL) approach, incorporating 1D convolutional neural network (1D CNN) blocks and hybrid features. These features included the top seven performing NLP-based features and 90 probabilistic features, allowing us to identify hidden patterns within these diverse features and improve the accuracy of our ACP prediction model. This is the first study to integrate spatial and probabilistic feature representations for predicting ACPs. Rigorous cross-validation and independent tests conclusively demonstrated that mACPpred 2.0 not only surpassed its predecessor (mACPpred) but also outperformed the existing state-of-the-art predictors, highlighting the importance of advanced feature representation capabilities attained through SDL. To facilitate widespread use and accessibility, we have developed a user-friendly for mACPpred 2.0, available at https://balalab-skku.org/mACPpred2/.

Asunto(s)

Antineoplásicos , Aprendizaje Profundo , Péptidos , Péptidos/química , Humanos , Antineoplásicos/farmacología , Biología Computacional/métodos , Programas Informáticos , Secuencia de Aminoácidos , Redes Neurales de la Computación

RESUMEN

Ripened pu-erh tea is known to have beneficial hypoglycemic properties. However, it remains unclear whether the bioactive peptides produced during fermentation are also related to hypoglycemic potential. This study aimed to identify hypoglycemic peptides in ripened pu-erh tea and to elucidate their bioactive mechanisms using physicochemical property prediction, molecular docking, molecular dynamics simulations, and cell experiments. Thirteen peptides were identified by liquid chromatography-mass spectrometry/mass spectrometry (LC-MS/MS). Among them, AADTDYRFS (AS-9) and AGDGTPYVR (AR-9) exhibited high α-glucosidase inhibitory activity, with half-maximal inhibitory concentration (IC50) values of 0.820 and 3.942 mg/mL, respectively. Molecular docking and dynamics simulations revealed that hydrogen bonding, hydrophobic interactions, and van der Waals forces assist peptides AS-9 and AR-9 in forming stable and tight complexes with α-glucosidase. An insulin-resistance (IR)-HepG2 cell model was established. AS-9 was non-toxic to IR-HepG2 cells and significantly increased the glucose consumption capacity, hexokinase, and pyruvate kinase activities of IR-HepG2 cells (p < 0.05). AS-9 alleviated glucose metabolism disorders and ameliorated IR by activating the IRS-1/PI3K/Akt signaling pathway and increasing the expression levels of MDM2, IRS-1, Akt, PI3K, GLUT4, and GSK3ß genes. In addition, no hemolysis of mice red blood cells red blood cells occurred at concentrations below 1 mg/mL. This work first explored hypoglycemic peptides in ripened pu-erh tea, providing novel insights for enhancing its functional value.

Asunto(s)

Inhibidores de Glicósido Hidrolasas , Hipoglucemiantes , Simulación del Acoplamiento Molecular , Péptidos , Té , Hipoglucemiantes/farmacología , Hipoglucemiantes/química , Animales , Té/química , Humanos , Células Hep G2 , Péptidos/química , Péptidos/farmacología , Inhibidores de Glicósido Hidrolasas/farmacología , Inhibidores de Glicósido Hidrolasas/química , Ratones , Simulación de Dinámica Molecular , Resistencia a la Insulina , Transducción de Señal/efectos de los fármacos , Proteínas Sustrato del Receptor de Insulina/metabolismo , Espectrometría de Masas en Tándem , alfa-Glucosidasas/metabolismo , Fermentación

RESUMEN

Peptides are valuable for therapeutic development, with multicyclic peptides showing promise in mimicking antigen-binding potency of antibodies. However, our capability to engineer multicyclic peptide scaffolds, particularly for the construction of large combinatorial libraries, is still limited. Here, we study the interplay of disulfide pairing between three biscysteine motifs, and designed a range of triscysteine motifs with unique disulfide-directing capability for regulating the oxidative folding of multicyclic peptides. We demonstrate that incorporating these motifs into random sequences allows the design of disulfide-directed multicyclic peptide (DDMP) libraries with up to four disulfide bonds, which have been applied for the successful discovery of peptide binders with nanomolar affinity to several challenging targets. This study encourages the use of more diverse disulfide-directing motifs for creating multicyclic peptide libraries and opens an avenue for discovering functional peptides in sequence and structural space beyond existing peptide scaffolds, potentially advancing the field of peptide drug discovery.

Asunto(s)

Cisteína , Disulfuros , Biblioteca de Péptidos , Disulfuros/química , Cisteína/química , Secuencias de Aminoácidos , Descubrimiento de Drogas/métodos , Secuencia de Aminoácidos , Péptidos/química , Péptidos/metabolismo , Péptidos Cíclicos/química , Péptidos Cíclicos/metabolismo , Unión Proteica , Humanos , Oxidación-Reducción , Pliegue de Proteína

RESUMEN

Binding-activated optical sensors are powerful tools for imaging, diagnostics, and biomolecular sensing. However, biosensor discovery is slow and requires tedious steps in rational design, screening, and characterization. Here we report on a platform that streamlines biosensor discovery and unlocks directed nanosensor evolution through genetically encodable fluorogenic amino acids (FgAAs). Building on the classical knowledge-based semisynthetic approach, we engineer ~15 kDa nanosensors that recognize specific proteins, peptides, and small molecules with up to 100-fold fluorescence increases and subsecond kinetics, allowing real-time and wash-free target sensing and live-cell bioimaging. An optimized genetic code expansion chemistry with FgAAs further enables rapid (~3 h) ribosomal nanosensor discovery via the cell-free translation of hundreds of candidates in parallel and directed nanosensor evolution with improved variant-specific sensitivities (up to ~250-fold) for SARS-CoV-2 antigens. Altogether, this platform could accelerate the discovery of fluorogenic nanosensors and pave the way to modify proteins with other non-standard functionalities for diverse applications.

Asunto(s)

Aminoácidos , Técnicas Biosensibles , Colorantes Fluorescentes , SARS-CoV-2 , Técnicas Biosensibles/métodos , Colorantes Fluorescentes/química , Humanos , SARS-CoV-2/genética , COVID-19/virología , Nanotecnología/métodos , Péptidos/metabolismo , Péptidos/química , Péptidos/genética

RESUMEN

Microcarriers for large-scale cell culture have a broader prospect in cell screening compared with the traditional high cost, low efficiency, and cell damaging methods. However, the equal biological affinity to cells has hindered its application. Therefore, based on the antifouling strategy of zwitterionic polymer, we developed a cell-specific microcarrier (CSMC) for shielding non-target cells and capturing mesenchymal stem cells (MSCs), which has characteristics of high biocompatibility, low background noise and high precision. Briefly, [2-(methacryloyloxy) ethyl] dimethyl-(3-sulfopropyl) ammonium hydroxide and glycidyl methacrylate were grafted onto polygalacturonic acid, respectively. The former built a hydration layer through solvation to provide an excellent antifouling surface, while the latter provided active sites for the click reaction with sulfhydryl-modified cell-specific peptides, resulting in rapid immobilization of peptides. This method is applicable to the vast majority of polysaccharide materials. The accurate capture ratio of MSCs by CSMC in a mixed multicellular environment is >95 % and the proliferation rate of MSCs on microcarriers is satisfactory. In summary, this grafting strategy of bioactive components lays a foundation for the application of polysaccharide materials in the biomedical field, and the specific adhesive microcarriers also open up new ideas for the development of stem cell screening as well.

Asunto(s)

Células Madre Mesenquimatosas , Pectinas , Péptidos , Células Madre Mesenquimatosas/citología , Pectinas/química , Péptidos/química , Metacrilatos/química , Proliferación Celular/efectos de los fármacos , Compuestos Epoxi/química , Humanos , Animales , Materiales Biocompatibles/química

RESUMEN

The potato has recently attracted more attention as a promising protein source. Potato proteins are commonly extracted from potato fruit juice, a byproduct of starch production. Potato proteins are characterized by superior techno-functional properties, such as water solubility, gel-forming, emulsifying, and foaming properties. However, commercially isolated potato proteins are often denatured, leading to a loss of these functionalities. Extensive research has explored the influence of different conditions and techniques on the emulsifying capacity and stability of potato proteins. However, there has been no comprehensive review of this topic yet. This paper aims to provide an in-depth overview of current research progress on the emulsifying capacity and stability of potato proteins and peptides, discussing research challenges and future perspectives. This paper discusses genetic diversity in potato proteins and various methods for extracting proteins from potatoes, including thermal and acid precipitation, salt precipitation, organic solvent precipitation, carboxymethyl cellulose complexation, chromatography, and membrane technology. It also covers enzymatic hydrolysis for producing potato-derived peptides and methods for identifying potato protein-derived emulsifying peptides. Furthermore, it reviews the influence of factors, such as physicochemical properties, environmental conditions, and food-processing techniques on the emulsifying capacity and stability of potato proteins and their derived peptides. Finally, it highlights chemical modifications, such as acylation, succinylation, phosphorylation, and glycation to enhance emulsifying capacity and stability. This review provides insight into future research directions for utilizing potato proteins as sustainable protein sources and high-value food emulsifiers, thereby contributing to adding value to the potato processing industry.

Asunto(s)

Péptidos , Proteínas de Plantas , Solanum tuberosum , Solanum tuberosum/química , Proteínas de Plantas/química , Péptidos/química , Emulsionantes/química , Emulsiones/química , Manipulación de Alimentos/métodos , Estabilidad Proteica

RESUMEN

RATIONALE: Elastin-like polypeptides (ELPs) are elastic and thermoresponsive biopolymers composed of VPGXG repeats (X can be any amino acid except proline), used in biomedical applications, for example, tissue engineering and drug delivery. As different variants of ELP are mostly produced fermentatively, there is a need for the development of analysis methods that allow for absolute protein quantification in both complex matrices and purified samples and MW determination of the final products. METHODS: ELPs were intracellularly expressed in Escherichia coli quantified after cell lysis and enzymatic digestion using a proline-specific protease ProAlanase (Promega) at acidic conditions. Resulting peptides were separated by liquid chromatography, and mass spectrometry analysis was conducted by electrospray ionization high-resolution mass spectrometry using an Orbitrap mass spectrometer. The addition of a stable isotopically labeled internal standard enabled quantification in complex matrices. Prior to intact mass analysis, ELPs were purified from fermentation broth by inverse temperature cycling. Intact protein analysis was performed using reversed-phase liquid chromatography, and mass spectrometry analysis was conducted by electrospray ionization high-resolution mass spectrometry using a time-of-flight mass spectrometer. RESULTS: Absolute quantification of ELPs was achieved by utilizing ELP-specific properties, that is, proline-rich, soluble at low pH and low temperature. The repetitive nature of ELPs allows for sensitivity increase and use of higher dilution factors to minimize the matrix effects. Despite the lack of amino acids with charged side chains (Arg, His, Lys, Asp, and Glu) in ELP, we demonstrated successful intact protein analysis using reversed-phase LC coupled to electrospray ionization TOF MS. Moreover, truncated protein forms could be chromatographically separated and characterized as well as N-terminal modifications. CONCLUSIONS: Both methods combined enabled quantitative and qualitative characterization of fermentatively produced ELPs.

Asunto(s)

Elastina , Escherichia coli , Péptidos , Elastina/química , Escherichia coli/química , Péptidos/química , Péptidos/análisis , Concentración de Iones de Hidrógeno , Espectrometría de Masa por Ionización de Electrospray/métodos , Frío , Cromatografía Liquida/métodos , Cromatografía de Fase Inversa/métodos , Polipéptidos Similares a Elastina

RESUMEN

The yeast pre1-1(ß4-S142F) mutant accumulates late 20S proteasome core particle precursor complexes (late-PCs). We report a 2.1 Å cryo-EM structure of this intermediate with full-length Ump1 trapped inside, and Pba1-Pba2 attached to the α-ring surfaces. The structure discloses intimate interactions of Ump1 with ß2- and ß5-propeptides, which together fill most of the antechambers between the α- and ß-rings. The ß5-propeptide is unprocessed and separates Ump1 from ß6 and ß7. The ß2-propeptide is disconnected from the subunit by autocatalytic processing and localizes between Ump1 and ß3. A comparison of different proteasome maturation states reveals that maturation goes along with global conformational changes in the rings, initiated by structuring of the proteolytic sites and their autocatalytic activation. In the pre1-1 strain, ß2 is activated first enabling processing of ß1-, ß6-, and ß7-propeptides. Subsequent maturation of ß5 and ß1 precedes degradation of Ump1, tightening of the complex, and finally release of Pba1-Pba2.

Asunto(s)

Microscopía por Crioelectrón , Complejo de la Endopetidasa Proteasomal , Proteínas de Saccharomyces cerevisiae , Saccharomyces cerevisiae , Complejo de la Endopetidasa Proteasomal/metabolismo , Proteínas de Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiae/química , Proteínas de Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo , Subunidades de Proteína/metabolismo , Subunidades de Proteína/química , Modelos Moleculares , Conformación Proteica , Péptidos/metabolismo , Péptidos/química , Unión Proteica , Chaperonas Moleculares

RESUMEN

The poor prognosis of hepatocellular carcinoma (HCC) is still an urgent challenge to be solved worldwide. Hence, assembling drugs and targeted short peptides together to construct a novel medicine delivery strategy is crucial for targeted and synergy therapy of HCC. Herein, a high-efficiency nanomedicine delivery strategy has been constructed by combining graphdiyne oxide (GDYO) as a drug-loaded platform, specific peptide (SP94-PEG) as a spear to target HCC cells, sorafenib, doxorubicin-Fe2+ (DOX-Fe2+), and siRNA (SLC7A11-i) as weapons to exert a three-path synergistic attack against HCC cells. In this work, SP94-PEG and GDYO form nanosheets with HCC-targeting properties, the chemotherapeutic drug DOX linked to ferrous ions increases the free iron pool in HCC cells and synergizes with sorafenib to induce cell ferroptosis. As a key gene of ferroptosis, interference with the expression of SLC7A11 makes the ferroptosis effect in HCC cells easier, stronger, and more durable. Through gene interference, drug synergy, and short peptide targeting, the toxic side effects of chemotherapy drugs are reduced. The multifunctional nanomedicine GDYO@SP94/DOX-Fe2+/sorafenib/SLC7A11-i (MNMG) possesses the advantages of strong targeting, good stability, the ability to continuously induce tumor cell ferroptosis and has potential clinical application value, which is different from traditional drugs.

Asunto(s)

Carcinoma Hepatocelular , Doxorrubicina , Ferroptosis , Neoplasias Hepáticas , Nanomedicina , Péptidos , Sorafenib , Ferroptosis/efectos de los fármacos , Carcinoma Hepatocelular/tratamiento farmacológico , Humanos , Neoplasias Hepáticas/tratamiento farmacológico , Doxorrubicina/farmacología , Doxorrubicina/química , Nanomedicina/métodos , Sorafenib/farmacología , Sorafenib/química , Línea Celular Tumoral , Animales , Péptidos/química , Péptidos/farmacología , Ratones , Antineoplásicos/farmacología , Antineoplásicos/química , Sinergismo Farmacológico , Sistema de Transporte de Aminoácidos y+/metabolismo , Ratones Desnudos , ARN Interferente Pequeño , Ratones Endogámicos BALB C , Sistemas de Liberación de Medicamentos/métodos

RESUMEN

Electrospray ionization is widely used to generate vapor phase ions for analysis by mass spectrometry in proteomics research. However, only a small fraction of the analyte enters the mass spectrometer due to losses that are fundamentally linked to the use of a background gas to stimulate the generation of ions from electrosprayed droplets. Here we report a nanopore ion source that delivers ions directly into high vacuum from aqueous solutions. The ion source comprises a pulled quartz pipette with a sub-100 nm opening. Ions escape an electrified meniscus by ion evaporation and travel along collisionless trajectories to the ion detector. We measure mass spectra of 16 different amino acid ions, post-translationally modified variants of glutathione, and the peptide angiotensin II, showing that these analytes can be emitted as desolvated ions. The emitted current is composed of ions rather than charged droplets, and more than 90% of the current can be recovered in a distant collector.

Asunto(s)

Aminoácidos , Iones , Nanoporos , Péptidos , Espectrometría de Masa por Ionización de Electrospray , Vacio , Aminoácidos/química , Péptidos/química , Espectrometría de Masa por Ionización de Electrospray/métodos , Proteómica/métodos , Angiotensina II/química

RESUMEN

The plasma membrane protein caveolin-1 (CAV-1) regulates signaling by inhibiting a wide range of kinases and other enzymes. Our previous study demonstrated that the downregulation of CAV-1 in psoriatic epidermal cells contributes to inflammation by enhancing JAK/STAT signaling, cell proliferation, and chemokine production. Administration of the CAV-1 scaffolding domain (CSD) peptide suppressed imiquimod (IMQ)-induced psoriasis-like dermatitis. To identify an optimal therapeutic peptide derived from CAV-1, we have compared the efficacy of CSD and subregions of CSD that have been modified to make them water soluble. We refer to these modified peptides as sCSD, sA, sB, and sC. In IMQ-induced psoriasis-like dermatitis, while all four peptides showed major beneficial effects, sB caused the most significant improvements of skin phenotype and number of infiltrating cells, comparable or superior to the effects of sCSD. Phosphorylation of STAT3 was also inhibited by sB. Furthermore, sB suppressed angiogenesis both in vivo in the dermis of IMQ-induced psoriasis mice and in vitro by blocking the ability of conditioned media derived from CAV-1-silenced keratinocytes to inhibit tube formation by HUVEC. In conclusion, sB had similar or greater beneficial effects than sCSD not only by cytokine suppression but by angiogenesis inhibition adding to its ability to target psoriatic inflammation.

Asunto(s)

Caveolina 1 , Citocinas , Imiquimod , Neovascularización Patológica , Psoriasis , Factor de Transcripción STAT3 , Psoriasis/tratamiento farmacológico , Psoriasis/inducido químicamente , Psoriasis/patología , Psoriasis/metabolismo , Caveolina 1/metabolismo , Animales , Ratones , Citocinas/metabolismo , Humanos , Factor de Transcripción STAT3/metabolismo , Neovascularización Patológica/tratamiento farmacológico , Neovascularización Patológica/metabolismo , Péptidos/farmacología , Péptidos/química , Piel/efectos de los fármacos , Piel/metabolismo , Piel/patología , Queratinocitos/efectos de los fármacos , Queratinocitos/metabolismo , Modelos Animales de Enfermedad , Agua/química , Solubilidad , Células Endoteliales de la Vena Umbilical Humana/efectos de los fármacos , Angiogénesis

RESUMEN

The Ebola delta peptide is an amphipathic, 40-residue peptide encoded by the Ebola virus, referred to as E40. The membrane-permeabilising activity of the E40 delta peptide has been demonstrated in cells and lipid vesicles suggesting the E40 delta peptide likely acts as a viroporin. The lytic activity of the peptide increases in the presence of anionic lipids and a disulphide bond in the C-terminal part of the peptide. Previous in silico work predicts the peptide to show a partially helical structure, but there is no experimental information on the structure of E40. Here, we use circular dichroism spectroscopy to report the secondary structure propensities of the reduced and oxidised forms of the E40 peptide in water, detergent micelles, and lipid vesicles composed of neutral and anionic lipids (POPC and POPG, respectively). Results indicate that the peptide is predominately a random coil in solution, and the disulphide bond has a small but measurable effect on peptide conformation. Secondary structure analysis shows large uncertainties and dependence on the reference data set and, in our system, cannot be used to accurately determine the secondary structure motifs of the peptide in membrane environments. Nevertheless, the spectra can be used to assess the relative changes in secondary structure propensities of the peptide depending on the solvent environment and disulphide bond. In POPC-POPG vesicles, the peptide transitions from a random coil towards a more structured conformation, which is even more pronounced in negatively charged SDS micelles. In vesicles, the effect depends on the peptide-lipid ratio, likely resulting from vesicle surface saturation. Further experiments with zwitterionic POPC vesicles and DPC micelles show that both curvature and negatively charged lipids can induce a change in conformation, with the two effects being cumulative. Electrostatic screening from Na+ ions reduced this effect. The oxidised form of the peptide shows a slightly lower propensity for secondary structure and retains a more random coil conformation even in the presence of PG-PC vesicles.

Asunto(s)

Dicroismo Circular , Ebolavirus , Micelas , Estructura Secundaria de Proteína , Ebolavirus/química , Fosfatidilcolinas/química , Soluciones , Fosfatidilgliceroles/química , Péptidos/química , Agua/química , Proteínas Virales/química , Secuencia de Aminoácidos

RESUMEN

An important functionality of lifelike "synthetic cells" is to mimic cell division. Currently, specialized proteins that induce membrane fission in living cells are the primary candidates for dividing synthetic cells. However, interactions between lipid membranes and proteins that are not found in living cells may also be suitable. Here, we discuss the potential of short membrane-anchored peptides to induce cell division. Specifically, we used the coarse-grained MARTINI model to investigate the interaction between short membrane-anchored peptides and a lipid bilayer patch. The simulation revealed that the anchored peptide induces significant spontaneous curvature and suggests that the lipid-peptide complex can be considered as a conically shaped "bulky headgroup" lipid. By systematically increasing the electrostatic charge of the peptide, we find that membrane-anchored peptides may generate sufficiently large constriction forces even at dilute coverages. Finally, we show that when the peptide has an opposite charge to the membrane, the peptide may induce division by binding the inner membrane leaflet of a synthetic cell, that is, cell division from within.

Asunto(s)

Membrana Dobles de Lípidos , Simulación de Dinámica Molecular , Péptidos , Membrana Dobles de Lípidos/química , Membrana Dobles de Lípidos/metabolismo , Péptidos/química , Péptidos/metabolismo , División Celular/efectos de los fármacos , Membrana Celular/química , Membrana Celular/metabolismo , Electricidad Estática

RESUMEN

Quinones are among the most important components in natural organic matter (NOM) for redox reactions; however, no quinones in complex environmental media have been identified. To aid the identification of quinone-containing molecules in ultracomplex environmental samples, we developed a chemical tagging method that makes use of a Michael addition reaction between quinones and thiols (-SH) in cysteine (Cys) and cysteine-contained peptides (CCP). After the tagging, candidates of quinones in representative aqueous environmental samples (water extractions of biochar) were identified through high-resolution mass spectrometry (HRMS) analysis. The MS and UV spectra analysis showed rapid reactions between Cys/CCP and model quinones with ß-carbon from the same benzene ring available for Michael addition. The tagging efficiency was not influenced by other co-occurring nonquinone representative compounds, including caffeic acid, cinnamic acid, and coumaric acid. Cys and CCP were used to tag quinones in water extractions of biochars, and possible candidates of quinones (20 and 53 based on tagging with Cys and CCP, respectively) were identified based on the HRMS features for products of reactions with Cys/CCP. This study has successfully demonstrated that such a Michael addition reaction can be used to tag quinones in complex environmental media and potentially determine their identities. The method will enable an in-depth understanding of the redox chemistry of NOM and its critical chemical compositions and structures.

Asunto(s)

Cisteína , Espectrometría de Masas , Péptidos , Quinonas , Cisteína/química , Péptidos/química , Quinonas/química , Carbón Orgánico/química