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

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

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

Probing the Binding Interfaces of Histone-Aptamer by Photo Cross-Linking Mass Spectrometry

Histone proteins, which could interact with DNA, play important roles in the regulation of chromatin structures, transcription, and other DNA-based biological processes. Here, we developed a novel aptamer-based probe for the analysis of histone H4-aptamer interfaces. This probe contains a DNA sequence for specific recognition of histone H4, a biotin tag for affinity enrichment, an aryl azide photoactive group for cross-linking and a cleavable disulfide group to dissociate aptamer from labeled histones. We successfully achieved specific enrichment of histone H4 and further developed a new analysis strategy for histone-aptamer interaction by photo cross-linking mass spectrometry. The binding area of histone H4 to aptamer was investigated and discussed for the first time. This strategy exhibits great potential and might further contribute to the understanding of histone–DNA interaction patterns

Maltose-Functionalized Hydrophilic Magnetic Nanoparticles with Polymer Brushes for Highly Selective Enrichment of N‑Linked Glycopeptides

Efficient enrichment glycoproteins/glycopeptides from complex biological solutions are very important in the biomedical sciences, in particular biomarker research. In this work, the high hydrophilic polyethylenimine conjugated polymaltose polymer brushes functionalized magnetic Fe₃O₄ nanoparticles (NPs) denoted as Fe₃O₄–PEI–pMaltose were designed and synthesized via a simple two-step modification. The obtained superhydrophilic Fe₃O₄–PEI–pMaltose NPs displayed outstanding advantages in the enrichment of N-linked glycopeptides, including high selectivity (1:100, mass ratios of HRP and bovine serum albumin (BSA) digest), low detection limit (10 fmol), large binding capacity (200 mg/g), and high enrichment recovery (above 85%). The above-mentioned excellent performance of novel Fe₃O₄–PEI–pMaltose NPs was attributed to graft of maltose polymer brushes and efficient assembly strategy. Moreover, Fe₃O₄–PEI–pMaltose NPs were further utilized to selectively enrich glycopeptides from human renal mesangial cell (HRMC, 200 μg) tryptic digest, and 449 N-linked glycopeptides, representing 323 different glycoproteins and 476 glycosylation sites, were identified. It was expected that the as-synthesized Fe₃O₄–PEI–pMaltose NPs, possessing excellent performance (high binding capacity, good selectivity, low detection limit, high enrichment recovery, and easy magnetic separation) coupled to a facile preparation procedure, have a huge potential in N-glycosylation proteome analysis of complex biological samples

Image_1_Four novel mutations identification in 17 beta-hydroxysteroid dehydrogenase-3 deficiency and our clinical experience: possible benefits of early treatment.tif

IntroductionIndividuals with 17-beta-hydroxysteroid dehydrogenase type 3 (17β-HSD3) deficiency face a multitude of challenges, primarily concerning genital appearance, potential malignancy risks, and fertility issues. This study reports our findings from an investigation involving five individuals affected by 17β-HSD3 deficiency, ranging in age from pre-adolescence to adolescence. Notably, we identified four previously unreported mutations in these subjects.MethodsOur study included a comprehensive evaluation to determine the potential occurrence of testicular tumors. The methods involved clinical examinations, genetic testing, hormone profiling, and patient history assessments. We closely monitored the progress of the study subjects throughout their treatment.ResultsThe results of this evaluation conclusively ruled out the presence of testicular tumors among our study subjects. Moreover, four of these individuals successfully underwent gender transition. Furthermore, we observed significant improvements in genital appearance following testosterone treatment, particularly among patients in the younger age groups who received appropriate treatment interventions.DiscussionThese findings underscore the critical importance of early intervention in addressing concerns related to genital appearance, based on our extensive clinical experience and assessments. In summary, our study provides insights into the clinical aspects of 17β-HSD3 deficiency, emphasizing the vital significance of early intervention in addressing genital appearance concerns. This recommendation is supported by our comprehensive clinical assessments and experience.</p

Image_2_Four novel mutations identification in 17 beta-hydroxysteroid dehydrogenase-3 deficiency and our clinical experience: possible benefits of early treatment.tif

IntroductionIndividuals with 17-beta-hydroxysteroid dehydrogenase type 3 (17β-HSD3) deficiency face a multitude of challenges, primarily concerning genital appearance, potential malignancy risks, and fertility issues. This study reports our findings from an investigation involving five individuals affected by 17β-HSD3 deficiency, ranging in age from pre-adolescence to adolescence. Notably, we identified four previously unreported mutations in these subjects.MethodsOur study included a comprehensive evaluation to determine the potential occurrence of testicular tumors. The methods involved clinical examinations, genetic testing, hormone profiling, and patient history assessments. We closely monitored the progress of the study subjects throughout their treatment.ResultsThe results of this evaluation conclusively ruled out the presence of testicular tumors among our study subjects. Moreover, four of these individuals successfully underwent gender transition. Furthermore, we observed significant improvements in genital appearance following testosterone treatment, particularly among patients in the younger age groups who received appropriate treatment interventions.DiscussionThese findings underscore the critical importance of early intervention in addressing concerns related to genital appearance, based on our extensive clinical experience and assessments. In summary, our study provides insights into the clinical aspects of 17β-HSD3 deficiency, emphasizing the vital significance of early intervention in addressing genital appearance concerns. This recommendation is supported by our comprehensive clinical assessments and experience.</p

Table_1_Four novel mutations identification in 17 beta-hydroxysteroid dehydrogenase-3 deficiency and our clinical experience: possible benefits of early treatment.docx

IntroductionIndividuals with 17-beta-hydroxysteroid dehydrogenase type 3 (17β-HSD3) deficiency face a multitude of challenges, primarily concerning genital appearance, potential malignancy risks, and fertility issues. This study reports our findings from an investigation involving five individuals affected by 17β-HSD3 deficiency, ranging in age from pre-adolescence to adolescence. Notably, we identified four previously unreported mutations in these subjects.MethodsOur study included a comprehensive evaluation to determine the potential occurrence of testicular tumors. The methods involved clinical examinations, genetic testing, hormone profiling, and patient history assessments. We closely monitored the progress of the study subjects throughout their treatment.ResultsThe results of this evaluation conclusively ruled out the presence of testicular tumors among our study subjects. Moreover, four of these individuals successfully underwent gender transition. Furthermore, we observed significant improvements in genital appearance following testosterone treatment, particularly among patients in the younger age groups who received appropriate treatment interventions.DiscussionThese findings underscore the critical importance of early intervention in addressing concerns related to genital appearance, based on our extensive clinical experience and assessments. In summary, our study provides insights into the clinical aspects of 17β-HSD3 deficiency, emphasizing the vital significance of early intervention in addressing genital appearance concerns. This recommendation is supported by our comprehensive clinical assessments and experience.</p