Adenosine Phosphate Functionalized Magnetic Mesoporous Graphene Oxide Nanocomposite for Highly Selective Enrichment of Phosphopeptides

Developing an efficient strategy to enrich the low abundance phosphopeptides before mass spectrometry detection is a vital preprocessing step in phosphoproteomics. In this work, we synthesized an adenosine phosphate-Ti⁴⁺ functionalized magnetic mesoporous graphene oxide nanocomposite (denoted as MG@mSiO₂-ATP-Ti⁴⁺) to selectively extract phosphorylated peptides from complex biological samples based on the immobilized metal ion affinity chromatography (IMAC). Mesoporous silica was coated on the substrate material of magnetic graphene oxide and then the ATP containing three phosphate groups was grafted on the inwall of mesoporous channels as chelating ligands to immobilize the Ti⁴⁺ cations. With favorable properties, such as large surface area and good hydrophilicity and size-exclusion effect, the MG@mSiO₂-ATP-Ti⁴⁺ exhibited excellent sensitivity and selectivity toward phosphopeptides whether in low concentration of β-casein digest (20 amol μL^–1, 4 fmol) or the digest mixture of β-casein and bovine serum albumin (with molar ratio of 1:1000) as well as good reusability. Furthermore, MG@mSiO₂-ATP-Ti⁴⁺ could also be applied in the selective enrichment of phosphorylated peptides from nonfat milk digest and human saliva and serum

Turn-on Fluorescent Sensing of Glutathione <i>S</i>‑Transferase at near-Infrared Region Based on FRET between Gold Nanoclusters and Gold Nanorods

A fluorescence resonance energy transfer (FRET) method based on gold nanoclusters capped glutathione (AuNCs@GSH) and amine-terminated gold nanorods (AuNRs) is designed for turn-on and near-infrared region (NIR) sensing of glutathione <i>S</i>-transferase (GST). The absorption band of AuNRs is tuned carefully to maximize the spectra overlap and enhance the efficiency of FRET. The FRET from multiple AuNCs to single AuNR quenches about 70% fluorescence emission of AuNCs. After GST is added, the strong specific interaction of GSH–GST can replace the AuNCs@GSH from AuNRs, FRET based on electrostatic interaction between AuNCs@GSH and AuNRs is switched off. Thus, emission enhancement of AuNCs@GSH is observed. The fluorescent enhancement is linearly with the increasing GST concentration over the range of 2–100 nM GST and the limit of detection for GST is about 1.5 nM

Multiepitope Templates Imprinted Particles for the Simultaneous Capture of Various Target Proteins

To achieve the simultaneous capture of various target proteins, the multiepitope templates imprinted particles were developed by phase inversion-based poly­(ether sulfone) (PES) self-assembly. Herein, with the top three high-abundance proteins in the human plasma, serum albumin, immunoglobulin G, and transferrin, as the target proteins, their N-terminal peptides were synthesized as the epitope templates. After the preorganization of three epitopes and PES in dimethylacetamide, the multiepitope templates imprinted particles were formed in water through self-assembly, by which the simultaneous recognition of three target proteins in human plasma was achieved with high selectivity. Furthermore, the binding kinetics study proved that the adsorption mechanism in this imprinting system toward three epitope templates was the same as that on the single-epitope imprinting polymer. These results demonstrate that our proposed multiepitope templates imprinting strategy might open a new era of artificial antibodies to achieve the recognition of various targets simultaneously

Dual-Functionalized Magnetic Metal–Organic Framework for Highly Specific Enrichment of Phosphopeptides

The highly specific enrichment of phosphoproteins and phosphopeptides from intricate biological systems is the precondition of in-depth phosphoproteome research. Herein, a novel dual-functionalized magnetic zirconium-based metal–organic framework (MOF) denoted as DFMMOF, with the purpose of combining the affinity of immobilized metal ion affinity chromatography (IMAC) and metal oxide affinity chromatography (MOAC) has been successfully synthesized. The inherent Zr–O cluster of DFMMOF particles acted as MOAC and the immobilized titanium­(IV) ions served for IMAC. The obtained DFMMOF exhibited rapid magnetic separation (within 5 s), large surface area (237.9 m² g^–1), high binding capacity (100 mg g^–1), and good postenrichment recovery (84.8%). Thanks to the strong affinity, low detection sensitivity (5 fmol) and high selectivity (β-casein/BSA with a molar ratio of 1:1000) for phosphopeptide enrichment were obtained using DFMMOF as absorbent. Moreover, the effective identification of phosphopeptides from real samples (human serum and nonfat milk) further confirmed the immense potential of DFMMOF as a promising candidate for the detection and extraction of trace amounts of phosphorylated peptides in complex biosamples

Enhancing the Mass Spectrometry Sensitivity for Oligonucleotide Detection by Organic Vapor Assisted Electrospray

There are two challenges in oligonucleotide detection by liquid chromatography coupled with mass spectrometry (LC-MS), the serious ion suppression effects caused by ion-pair reagents and the low detection sensitivity in positive mode MS. In this study, highly concentrated alcohol vapors were introduced into an enclosed electrospray ionization chamber, and oligonucleotides could be well detected in negative mode MS even with 100 mM triethylammonium acetate (TEAA) as an ion-pair reagent. The MS signal intensity was improved 600-fold (for standard oligonucleotide dT15) by the isopropanol vapor assisted electrospray, and effective ion-pair LC separation was feasibly coupled with high-sensitive MS detection. Then, oligonucleotides were successfully detected in positive mode MS with few adducts by propanoic acid vapor assisted electrospray. The signal intensity was enhanced more than 10-fold on average compared with adding acids into the electrospray solution. Finally, oligonucleotides and peptides or histones were simultaneously detected in MS with little interference with each other. Our strategy provides a useful alternative for investigating the biological functions of oligonucleotides

The Current Model Of Communication Processes In Banks

Розглянуто основні складові сучасної моделі комунікаційних процесів та новітні підходи щодо їх реалізаціїThe basic components of a modern model of communication processes and new approaches to their implementatio

Clickable Periodic Mesoporous Organosilica Monolith for Highly Efficient Capillary Chromatographic Separation

A novel clickable periodic mesoporous organosilica monolith with the surface area up to 1707 m² g^–1 was in situ synthesized in the capillary by the one-step condensation of the organobridged-bonded alkoxysilane precursor bis­(triethoxysilyl)­ethylene. With Si–C bonds in the skeleton, the monolith possesses excellent chemical and mechanical stability. With vinyl groups highly loaded and homogeneously distributed throughout the structure, the monolith can be readily functionalized with functional groups by effective thiol–ene “click” chemistry reaction. Herein, with “click” modification of C18, the obtained monolith was successfully applied for capillary liquid chromatographic separation of small molecules and proteins. The column efficiency could reach 148 000 N/m, higher than most reported hybrid monoliths. Moreover, intact proteins could be separated well with good reproducibility, even after the monolithic column was exposed by basic mobile phase (pH 10.0) overnight, demonstrating the great promising of such monolith for capillary chromatographic separation

Click Synthesis of Hydrophilic Maltose-Functionalized Iron Oxide Magnetic Nanoparticles Based on Dopamine Anchors for Highly Selective Enrichment of Glycopeptides

The development of methods to isolate and enrich low-abundance glycopeptides from biological samples is crucial to glycoproteomics. Herein, we present an easy and one-step surface modification strategy to prepare hydrophilic maltose functionalized Fe₃O₄ nanoparticles (NPs). First, based on the chelation of the catechol ligand with iron atoms, azido-terminated dopamine (DA) derivative was assembled on the surface of magnetic Fe₃O₄ nanoparticles by sonication. Second, the hydrophilic maltose-functionalized Fe₃O₄ (Fe₃O₄-DA-Maltose) NPs were obtained via copper­(I)-catalyzed azide–alkyne cycloaddition (click chemistry). The morphology, structure, and composition of Fe₃O₄-DA-Maltose NPs were investigated by Fourier transform infrared spectroscopy (FT-IR), transmission electron microscopy (TEM), X-ray powder diffraction (XRD), X-ray photoelectron spectrometer (XPS), and vibrating sample magnetometer (VSM). Meanwhile, hydrophilicity of the obtained NPs was evaluated by water contact angle measurement. The hydrophilic Fe₃O₄-DA-Maltose NPs were applied in isolation and enrichment of glycopeptides from horseradish peroxidase (HRP), immunoglobulin (IgG) digests. The MALDI-TOF mass spectrometric analysis indicated that the novel NPs exhibited high detection sensitivity in enrichment from HRP digests at concentration as low as 0.05 ng μL^–1, a large binding capacity up to 43 mg g^–1, and good recovery for glycopeptides enrichment (85–110%). Moreover, the Fe₃O₄-DA-Maltose NPs were applied to enrich glycopeptides from human renal mesangial cells (HRMC) for identification of N-glycosylation sites. Finally, we identified 115 different N-linked glycopeptides, representing 93 gene products and 124 glycosylation sites in HRMC

Hydrophobic Tagging-Assisted N‑Termini Enrichment for In-Depth N‑Terminome Analysis

The analysis of protein N-termini is of great importance for understanding the protein function and elucidating the proteolytic processing. Herein, we develop a negative enrichment strategy, termed as hydrophobic tagging-assisted N-termini enrichment (HYTANE) to achieve a global N-terminome analysis. The HYTANE strategy showed a high efficiency in hydrophobic tagging and C18 material-assisted depletion using bovine serum albumin (BSA) as the sample. This strategy was applied to N-termini profiling from <i>S. cerevisiae</i> cell lysates and enabled the identification of 1096 protein N-termini, representing the largest N-terminome data set of <i>S. cerevisiae</i>. The identified N-terminal peptides accounted for 99% of all identified peptides, and no deficiency in acidic, histidine (His)-containing, and His-free N-terminal peptides was observed. The presented HYTANE strategy is therefore a highly selective, efficient, and unbiased strategy for the large scale N-terminome analysis. Furthermore, using the HYTANE strategy, we identified 329 cleavage sites and 291 substrates of caspases in Jurkat cells, demonstrating the great promise of HYTANE strategy for protease research. Data are available via ProteomeXchange with identifier PXD004690

Thermoresponsive Epitope Surface-Imprinted Nanoparticles for Specific Capture and Release of Target Protein from Human Plasma

Among various artificial antibodies, epitope imprinted polymer has been paid increasingly attention. To modulate the “adsorption and release” behavior by environment stimuli, <i>N</i>-isopropylacrylamide, was adopted to fabricate the thermoresponsive epitope imprinted sites. The prepared imprinted materials could adsorb 46.6 mg/g of target protein with the imprinting factor of 4.0. The template utilization efficiency could reach as high as 8.21%. More importantly, in the real sample, the materials could controllably capture the target protein from the human plasma at 45 °C and release it at 4 °C, which demonstrated the “on-demand” application potentials of such materials in the biomolecule recognition field