Velvet mesquite (Prosopis velutina) is a native legume of the southwestern United States and northwestern Mexico, contributing significantly to the desert ecosystem and playing key ecological roles. It is also an important cause of allergic respiratory disease widely distributed in the Sonoran, Chihuahuan, and Mojave Deserts. However, no allergens from velvet mesquite pollen have been identified to date. Pollen proteins were extracted and analyzed by one- and two-dimensional electrophoresis and immunoblotting using a pool of 11 sera from mesquite-sensitive patients as the primary antibody. IgE-recognized protein spots were identified by mass spectrometry and bioinformatics analysis. Twenty-four unique proteins, including proteins well known as pollen, food, airway, or contact allergens and four proteins not previously reported as pollen allergens, were identified. This is the first report on allergenic proteins in velvet mesquite pollen. These findings will contribute to the development of specific diagnosis and treatment of mesquite pollen allergy.
Nanomedicine has led to the development of new biocompatible and biodegradable materials able to improve the pharmaceutical effect of bioactive components, broadening the options of treatment for several diseases, including cancer. Additionally, some snake venom toxins have been reported to present cytotoxic activity in different tumor cell lines, making them an auspicious option to be used as cancer drugs. The present study aims to evaluate the cytotoxic activity of the northern black-tailed rattlesnake (Crotalus molossus molossus) venom-loaded chitosan nanoparticles (Cs-Venom NPs) against the T-47D breast carcinoma cell line. To do so, we first identified the significant proteins composing the venom; afterward, hemocompatibility and cytotoxic activity against tumoral cells were evaluated. The venom was then loaded into chitosan nanoparticles through the ionotropic gelation process, obtaining particles of 415.9±21.67 nm and ζ-potential of +28.3±1.17 mV. The Cs-Venom complex delivered the venom into the breast carcinoma cells, inhibiting their viability and inducing morphological changes in the T-47D cells. These features indicate that these nanoparticles are suitable for the potential use of C. m. molossus venom toxins entrapped within polymer nanoparticles for the future development and research of cancer drugs.
Antineoplastic Agents/pharmacology , Breast Neoplasms/pathology , Chitosan/chemistry , Crotalid Venoms/pharmacology , Nanoparticles/chemistry , Animals , Antineoplastic Agents/chemistry , Breast Neoplasms/drug therapy , Breast Neoplasms/metabolism , Cell Line, Tumor , Cell Survival/drug effects , Crotalid Venoms/chemistry , Crotalus , Drug Delivery Systems/methods , Female , Humans , Male , Nanomedicine/methods , Snake Venoms/pharmacology
In this work, previously synthesized and characterized core-shell silica nanoparticles (FCSNP) functionalized with immobilized molecular bait, Cibacron blue, and a porous polymeric bis-acrylamide shell were incubated with pooled urine samples from adult women or men with normal weight, overweight or obesity for the isolation of potential biomarkers. A total of 30 individuals (15 woman and 15 men) were included. FCSNP allowed the capture of a variety of low molecular weight (LMW) proteins as evidenced by mass spectrometry (MS) and the exclusion of high molecular weight (HMW) proteins (>34 kDa) as demonstrated by SDS-PAGE and 2D SDS-PAGE. A total of 36 proteins were successfully identified by MS and homology database searching against the Homo sapiens subset of the Swiss-Prot database. Identified proteins were grouped into different clusters according to their abundance patterns. Four proteins were found only in women and five only in men, whereas 27 proteins were in urine from both genders with different abundance patterns. Based on these results, this new approach represents an alternative tool for isolation and identification of urinary biomarkers.
Obesity/urine , Proteinuria/urine , Proteomics , Adult , Biomarkers/urine , Female , Humans , Male , Middle Aged
BACKGROUND: Red oak pollen is an important cause of allergic respiratory disease and it is widely distributed in North America and central Europe. To date, however, red oak pollen allergens have not been identified. Here, we describe the allergenic protein profile from red oak pollen. METHODS: Total proteins were extracted from red oak pollen using a modified phenolic extraction method, and, subsequently, proteins were separated by two-dimensional gel electrophoresis (2DE) for both total protein stain (Coomassie Blue) and immunoblotting. A pool of 8 sera from red oak sensitive patients was used to analyze blotted proteins. Protein spots were analyzed by Mass Spectrometry. RESULTS: Electrophoretic pattern of total soluble proteins showed higher intensity bands in the regions of 26-40 and 47-52 kDa. Two dimensional immunoblots using pool sera from patients revealed four allergenic proteins spots with molecular masses in the range from 50 to 55 kDa. Mass spectrometry analysis identified 8 proteins including Enolase 1 and Enolase 1 chloroplastic, Xylose isomerase (X1 isoform), mitochondrial Aldehyde dehydrogenase, UTP-Glusose-1-phosphate uridylyltransferase, Betaxylosidase/alpha-l-arabinofuranosidase and alpha- and beta subunits of ATP synthase. CONCLUSIONS: This study has identified for first time 8 IgE binding proteins from red oak pollen. These findings will pave the way towards the development of new diagnostic and therapeutic modalities for red oak allergy.
Probiotics are live microorganisms conferring health benefits when administered in adequate amounts. However, the passage through the gastrointestinal tract represents a challenge due to pH variations, proteases, and bile salts. This study aimed to evaluate the proteomic response of Saccharomyces boulardii to simulated gastrointestinal digestion and the influence of encapsulation on yeast viability. Different pH values and time periods simulating the passage through different sections of the gastrointestinal tract were applied to unencapsulated and encapsulated yeasts. Encapsulation in 0.5% calcium alginate did not improve yeast survival or induce changes in protein patterns whereas protein extracts from control and digested yeasts showed remarkable differences when separated by SDS-PAGE. Protein bands were analyzed by tandem mass spectrometry. Protein identification revealed unique proteins that changed acutely in abundance after simulated digestion. Carbohydrate metabolism, protein processing, and oxide-reduction were the biological processes most affected by simulated gastrointestinal digestion in S. boulardii.
Herein, silica nanoparticles were synthesized and chemically modified with iminodiacetic acid (IDA) and Ni2+ ions surrounded by a bis-acrylamide polymeric shell to obtain a new core-shell immobilized metal affinity chromatography (IMAC) based material. These Ni2+-IDA-core-shell silica nanoparticles (Ni2+-IDA-CSS-NP) represent a new alternative for purification of His-tagged proteins and exclusion of high molecular weight (HMW) proteins at the same time. Nanoparticles presented a final size of 479.6 ± 6.9 nm determined by dynamic light scattering (DLS) and a surface charge of -37.2 ± 0.5 mV. Successful incorporation of the different compounds at every phase of synthesis was evidenced by ATR-FTIR analysis. Ni2+-IDA-CSS-NP were used for isolation of His-tagged spo0F (6His-spo0F) from E. coli lysate. Ni2+-IDA-CSS-NP presented a capacity of 4.16 ± 0.45 µg mg-1. Purification of 6His-spo0F with high selectivity and the effective exclusion of HMW proteins were evidenced by SDS-PAGE and validated through mass spectrometry analysis.
Silica nanoparticles were functionalized with immobilized molecular bait, Cibacron Blue, and a porous polymeric bis-acrylamide shell. These nanoparticles represent a new alternative to capture low molecular weight (LMW) proteins/peptides, that might be potential biomarkers. Functionalized core-shell silica nanoparticles (FCSNP) presented a size distribution of 243.9 ± 11.6 nm and an estimated surface charge of -38.1 ± 0.9 mV. The successful attachment of compounds at every stage of synthesis was evidenced by ATR-FTIR. The capture of model peptides was determined by mass spectrometry, indicating that only the peptide with a long sequence of hydrophobic amino acids (alpha zein 34-mer) interacted with the molecular bait. FCSNP excluded the high molecular weight protein (HMW), BSA, and captured LMW proteins (myoglobin and aprotinin), as evidenced by SDS-PAGE. Functionalization of nanoparticles with Cibacron Blue was crucial to capture these molecules. FCSNP were stable after twelve months of storage and maintained a capacity of 3.1-3.4 µg/mg.
Nanoparticles/chemistry , Peptides/chemistry , Proteins/chemistry , Silicon Dioxide/chemistry , Adsorption , Chemistry Techniques, Synthetic , Hydrophobic and Hydrophilic Interactions , Molecular Weight , Nanoparticles/ultrastructure , Particle Size , Spectroscopy, Fourier Transform Infrared
The apple is a food rich in diverse classes of polyphenols (PP), among which the proanthocyanidins (PCs), which are primarily concentrated in the skin, are one of the most abundant. These compounds are of considerable interest for their possible positive health effects because of their antioxidant properties. However, depending on the classes of PP present (chemical composition) and their relative concentrations in the apple skin, their antioxidant effects vary and some of their components can even generate prooxidant effects. This work determined the chemical composition and antioxidant-prooxidant potential of a polyphenolic extract (PPE) and a proanthocyanidin-rich fraction (PRF) of apple skin, along with the contribution of their most abundant individual compounds, based on their copper chelating ability, ease in reducing peroxidase-generated free radicals and TEAC (Trolox-Equivalent Antioxidant Capacity) assay. For this purpose, chromatographic and colorimetric methods were used. The majority compounds identified in PPE were flavan-3-ols (44.58%), flavonols (42.89%) and dihydrochalcones (11.60%). In PRF, we detected monomers and oligomers from dimers to heptamers, which were composed of 97% (-)-epicatechin and 3% (+)-catechin. The antioxidant potential was notably higher in PRF than in PPE. The (-)-epicatechin monomer and the procyanidin B2 dimer showed more ease in reducing peroxidase-generated free radicals compared to other compounds of the apple skin, whereas phloridzin dihydrochalcone produced prooxidant effects.
