ANALYTICA CHIMICA ACTA
·
2018
·
IF 5.123
Haiyang Zhang,Xiaowei Li,Yating Yao,Shujuan Ma,Zhen Liu,Junjie Ou,Yinmao Wei,Mingliang Ye
Hierarchically porous monoliths as a key feature of biological materials have been applied in enrichment and separation. In this work, a metal immobilized hierarchically porous organosilica hybrid monolith was synthesized by hydrolysis and condensation of tetraethoxysilane (TEOS) and diethoxyphosphorylethyl-triethoxysilane (DPTS) under alkaline environment . Phosphonate ester groups were firstly introduced by the employment of DPTS as functional monomer , and then acidified to phosphonic acids . The surface area of optimal monolith could reach to 1170 m 2 /g, which simultaneously contained micropores and mesopores (4 nm) obtained from nitrogen sorption measurement. Meanwhile, mercury intrusion porosimetry (MIP) further demonstrated that macropores (1–3 μm) existed in monoliths. Followed by chelating with titanium ion (Ti 4+ ), the hierarchically porous organosilica hybrid monoliths could be applied as IMAC materials. This synthesized process was easy-operating and time-saving, and avoided the tedious and complex process of traditional Ti 4+ -IMAC materials. Furthermore, the Ti 4+ -IMAC monoliths exhibited high adsorption capacity for pyridoxal 5′-phosphate (82.6 mg/g). The 3282 unique phosphopeptides could be identified from 100 μg of HeLa digests after enrichment with the monolith, exhibiting excellent enrichment performance of low-abundance phosphopeptides.
研究领域:
生物材料
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分离科学
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分析化学
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DOI
PubMed
高影响因子
MOLECULES
·
2026
·Vol. 31
·期 3
·396
·
IF 4.6
Yan Lu,Sen Zhang,Hong-Yan Ge,Han-Yue Yang,Feng Zhang,Yi-Fan Pan,Hong-Zhen Lian
Protein phosphorylation is a crucial post-translational modification that regulates protein activity, cellular signaling, transcriptional regulation, and cell cycle control. However, the analysis of phosphoproteins in biological samples is often compromised by complex sample matrices and interference from high-abundance proteins. While the top-down phosphoproteomics strategy enables comprehensive analysis of post-translational modifications based on intact proteins, its requirement for higher protein purity due to low protein ionization efficiency poses stern challenges. Consequently, developing appropriate enrichment methods for phosphoproteins in practical samples becomes essential. Immobilized metal ion affinity chromatography (IMAC) represents a common strategy for phosphorylated protein separation and enrichment. Among metal ions, Ti4+has gained widespread application as IMAC chelating ligands due to its capacity to form multiple coordination networks and its high selectivity for phosphorylated protein enrichment, leveraging the strong chelating ability of phosphate groups toward metal ions. This paper presents the design and preparation of a novel magnetic Ti-IMAC nanocomposite, MNP@MPTMS–VPA–Ti(IV). The material is modified with phosphate groups via facile thiol-ene click chemistry and then immobilizes Ti4+, enabling selective enrichment of intact phosphoproteins through IMAC affinity. The efficiency of enrichment was evaluated using subsequent matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) for detection and analysis. This Ti-IMAC material-based magnetic solid-phase extraction (MSPE)-MALDI-TOF MS protocol has been successfully applied to enrich intact phosphoproteins in milk and eel mucus with high selectivity, sensitivity, and suitability.
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PubMed
高影响因子
ANALYST
·
2023
·Vol. 148
·期 14
·3392-3402
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IF 4.2
Danni Wang,Quanshou Feng,Yiting Luo,Weimin Wang,Yinghua Yan,Chuan-Fan Ding
In this work, titanium-rich hydrazide-linked porous organic polymers (hydrazide-POPs-Ti4+) were synthesized using hydrazine, 2,3-dihydroxyterephthalaldehyde (DHTA) and trimethyl 1,3,5-benzenetricarboxylate (TP) as the ligands. Hydrazide-POPs-Ti4+ combined with HILIC and IMAC can be used for simultaneous enrichment of glycopeptides and phosphopeptides. The detection limit of this protocol is 0.1 fmol μL−1 for glycopeptides and 0.005 fmol μL−1 for phosphopeptides, and the selectivities are 1 : 1000 and 1 : 2000 for glycopeptides and phosphopeptides, respectively. For practical bio-sample analysis, 201 glycopeptides associated with 129 glycoproteins and 26 phosphopeptides associated with 21 phosphoproteins were selectively captured from healthy human serum, and 186 glycopeptides associated with 117 glycoproteins and 60 phosphopeptides associated with 50 phosphoproteins were enriched in the serum of breast cancer patients. Gene Ontology analysis indicated that the identified glycoproteins and phosphoproteins were linked to breast cancer, including the binding of complement component C1q and low-density lipoprotein particles, protein oxidation and complement activation, suggesting that these connected pathways are probably engaged in the disease pathology of breast cancer.
研究领域:
分析化学
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生物化学
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分子生物学
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癌症研究
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DOI
PubMed
高影响因子
GREEN CHEMISTRY
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2021
·Vol. 24
·期 1
·238-250
·
IF 10.182
Lei Pan,Shujuan Ma,Ruizhi Tang,Wenrui Wu,Junjie Ou,Cong Li,Yehua Shen
Protein phosphorylation is involved in many biological processes and associated with some diseases. However, because of the low abundance and large dynamic changes of phosphopeptides in biological samples, it is necessary to enrich phosphopeptides before mass spectrometry detection. Many kinds of materials have been developed for phosphopeptide enrichment, but most of them are disposable and their preparation processes are toxic. In this study, a green strategy was presented to fabricate immobilized metal affinity chromatography (IMAC) materials. Using chitosan as a precursor, polyethylene glycol diglycidyl ether as a cross-linking agent and 2,3,4-trihydroxybenzaldehyde as a functional monomer, a series of chitosan membranes (CMs) with free-standing lamellar 3D architectures were prepared via a freeze-casting technique and after chelating with titanium ions (Ti4+) could serve as IMAC adsorbents for phosphopeptide enrichment in biological samples. Due to the stronger chelating force between Ti4+ and the pyrogallol ligand than that between Ti4+ and the phosphate groups, the IMAC material is reusable for the enrichment of phosphopeptides, exhibiting an excellent enrichment performance and a higher capacity than other IMAC materials such as organosilica hybrid monolith and carbonaceous spheres. Besides, four endogenous phosphopeptides and two dephosphorylated fragments were captured by Ti4+-CM from the human serum. Therefore, Ti4+-CM demonstrates potential commercialization prospects because of its green preparation process and reusability.
研究领域:
生物化学
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材料科学
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分析化学
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DOI
高影响因子
ANALYTICAL LETTERS
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2023
·
IF 2
Menglin Ouyang,Jiani Wu,Yinghua Yan,Chuan-Fan Ding
Inspired by the enrichment preferences of metal ions for phosphopeptides, a layered assembly method was adopted to introduce hydrophilic polyhydroxy ligands tannic acid (TA) to prepare magnetic bimetal ion-functionalized nanomaterials denoted by Fe3O4@TA@Ti4+/Zr4+ to provide the comprehensive enrichment of phosphopeptides. Benefiting from the excellent ion immobilization ability and unbiasedness for phosphopeptide enrichment, Fe3O4@TA@Ti4+/Zr4+ shows excellent properties for the enrichment and identification of global phosphopeptides. The results show that the material possesses high efficiency, outstanding selectivity (the maximum molar ratio β-casein to bovine serum albumin is 1:1000), a low detection limit of 0.4 fmol/μL, and the ability to be recycled at least 10 times. In addition, excellent performance was also demonstrated in the analysis of skimmed milk and human saliva: 13 and 31 phosphopeptides were successfully captured. The results demonstrate that the material has high potential for practical application in phosphoproteomics.
研究领域:
蛋白质组学
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分析化学
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生物医学
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DOI
高影响因子
TALANTA
·
2023
·
IF 6.1
Kexuan Wang,Ajuan Yu,Yu Gao,Miao Chen,Hang Yuan,Shusheng Zhang,Gangfeng Ouyang
A novel immobilized metal affinity chromatography (IMAC) functional composite, [email protected] @ [email protected] 4+ , was fabricated based on ultrathin magnetic nitrogen-doped graphene tube ( [email protected] ) after chelated Ti 4+ with polydopamine, following as a magnetic solid-phase extraction sorbent for rapidly selective enrichment and mass spectrometry identification of phosphorylated peptides. After optimized, the composite exhibited high specificity in the enrichment of phosphopeptides from the digest mixture of β -casein and bovine serum albumin (BSA). The robust method presented the low detection limits (1 fmol, 200 μL) and excellent selectivity (1:100) in the molar ration mixture of β -casein and BSA digests. Furthermore, the selective enrichment of phosphopeptides in the complex bio-samples, was successfully carried out. The results showed that 28 phosphopeptides were finally detected in mouse brain, and 2087 phosphorylated peptides were identified in the HeLa cells extracts with specific selectivity of 95.6%. The enrichment performance of [email protected] @ [email protected] 4+ was satisfactory, suggesting that the functional composite provided a potential application in the enrichment of trace phosphorylated peptides from the complex biological matrix.
研究领域:
分析化学
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生物化学
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蛋白质组学
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生物技术
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PubMed
高影响因子
BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS
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2024
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IF 2.5
Luyu Qi,Jun Zhu,Zhongyi Cheng,Zhiyi Yuan,Wulin Qi,JiaHuan Wu,Yifeng Qin,Jingbo Yang,Tao Luo,Minkun Wang,Yejing Weng,Jianzhong Shao
Protein phosphorylation, a widely occurring and significant post-translational modification, is integral to various biological processes. We previously utilized a protein affinity probe to identify genes damaged by cisplatin, revealing that it inflicts substantial damage on protein kinase and protein phosphatase genes. In this study, we investigated cisplatin-induced alterations in the global proteome and phosphoproteome of A549 cells. Employing Fe-IMAC beads and tyrosine phosphorylation enrichment antibodies, we identified 6944 protein groups and 18,274 phosphorylation sites on 4915 proteins across three biological replicates of both cisplatin-treated A549 cells and control cells. Among these, 730 tyrosine phosphorylation sites were identified—marking the most substantial discovery of such sites in A549 cells following cisplatin treatment. Bioinformatics analysis indicated that the proteins exhibiting significant phosphorylation level changes predominantly involved in RNA processing, modification, transcription, translation, and the spliceosome. This suggests that cisplatin-induced damage to protein kinases and phosphatases may disrupt the normal function of these proteins, consequently impairing DNA replication, RNA translation, and shearing, ultimately culminating in tumor cell death. Moreover, we cross-referenced our proteomic data with our previously obtained cisplatin-damaged genes, observing that the majority of down-regulated proteins derived from cisplatin-induced gene damage. The data are available on ProteomeXchange under the identifier PXD053902.
研究领域:
癌症生物学
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分子生物学
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蛋白质组学
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磷酸蛋白质组学
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生物信息学
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DOI
PubMed
高影响因子
ANALYST
·
2023
·Vol. 148
·期 19
·4738-4745
·
IF 4.2
Luyan Meng,Bing Wang,Baichun Wang,Quanshou Feng,Sijia Zhang,Zi Xiong,Shun Zhang,Ting Cai,Chuan-Fan Ding,Yinghua Yan
A Ti4+-functionalized magnetic covalent organic framework material with flexible branched polymers (mCOF@ε-PL@THBA-Ti4+) built via an immobilized metal ion affinity chromatography (IMAC) enrichment strategy was proposed through post-synthesis modification. Hydrophilic ε-poly-L-lysine (ε-PL) rich in amino active groups was first introduced in the fabrication of the phosphopeptide enrichment material to increase the hydrophilicity while providing more functional modification pathways of the material. 2,3,4-Trihydroxy-benzaldehyde (THBA) provides abundant binding sites for the immobilization of numerous Ti4+, which is advantageous for the subsequent efficient phosphopeptide enrichment. The magnetic nanocomposite exhibited outstanding performance of phosphopeptide enrichment with good selectivity (1 : 5000), a low detection limit (2 fmol), and relatively high loading capacity (66.7 mg g−1). What's more, after treatment with mCOF@ε-PL@THBA-Ti4+, 16 endogenous phosphopeptides from fresh saliva of healthy people were recognized by MALDI-TOF MS, and 50 phosphopeptides belonging to 35 phosphoproteins from the serum of uremia patients were detected by nano-LC-MS/MS. Proteomics data analysis for the differential protein selection between uremia and normal controls was conducted using R software, and four down-regulated and three up-regulated proteins were obtained. The results suggested that the prepared material has potential applications in biomarker discovery.
研究领域:
材料科学
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分析化学
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生物医学工程
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蛋白质组学
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DOI
PubMed
高影响因子
INTERNATIONAL JOURNAL OF BIOLOGICAL MACROMOLECULES
·
2024
·
IF 7.7
Xiaoya Zhang,Jiakai Chen,Xiuqin Sheng,Chuan-Fan Ding,Yinghua Yan
Chitosan, as a biological macromolecule with excellent biocompatibility, has great potential for application in immobilized metal affinity chromatography (IMAC) strategies. In-depth analysis of low-abundance phosphopeptides in organisms can help reveal the pathological mechanisms of diseases. Here, we developed an IMAC material based on a biomimetic honeycomb chitosan membrane. The material demonstrates excellent biocompatibility, good hydrophilicity, and strong metal chelating capacity, which collectively confer outstanding enrichment properties. The material has high sensitivity (0.05 fmol), great selectivity (1:2000), excellent cycling stability (at least 10 cycles) and acid-base stability. In addition, the material was employed in human serum, successfully enriching 129 phosphopeptides from the serum of gastric cancer patients and 146 phosphopeptides from healthy controls. Sequence logo suggests a potential association between gastric cancer and glutamine. Ultimately, an in-depth gene ontology analysis was carried out on the phosphopeptides that were enriched in the serum samples. Compared to normal controls, our results demonstrated dysregulated expression of biological process, cellular component, and molecular function in gastric cancer patients. This suggests that the disease involves, such as blood coagulation pathways, cholesterol metabolism, and heparin binding. All experimental outcomes converge to demonstrate the substantial promise of the material for applications within proteomics research.
研究领域:
生物技术
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蛋白质组学
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癌症研究
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材料科学
·
生物化学
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PubMed
高影响因子
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