A novel IMAC platform – adenosine coupled functional magnetic microspheres for phosphoproteome research

Yan, Yinghua; Lu, Yujie; Chen, Mengying; Liang, Hongze

doi:10.1039/c7ay02931d

Cited by 4 publications

(2 citation statements)

References 40 publications

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“…ATP has three phosphate groups that can chelate heavy metal ions, which in turn can bind to phosphopeptides. Previous studies have used ATP as a ligand in IMAC materials and achieved excellent selectivity and sensitivity in phosphopeptide enrichment. − However, the strong hydrophilicity of ATP provided by its hydrophilic adenine base and ribose sugar groups has not been exploited, which can superimpose the HILIC-mode enrichment upon the electrostatic attraction. The structure and good biocompatibility of ATP make it a promising ligand for the purpose.…”

Section: Introductionmentioning

confidence: 99%

ATP-Coated Dual-Functionalized Titanium(IV) IMAC Material for Simultaneous Enrichment and Separation of Glycopeptides and Phosphopeptides

Wang,

Huang,

Zhang

et al. 2023

J. Proteome Res.

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Protein glycosylation and phosphorylation are two of the most common post-translational modifications (PTMs), which play an important role in many biological processes. However, low abundance and poor ionization efficiency of phosphopeptides and glycopeptides make direct MS analysis challenging. In this study, we developed a hydrophilicity-enhanced bifunctional Ti-IMAC (IMAC: immobilized metal affinity chromatography) material with grafted adenosine triphosphate (denoted as epoxy-ATP-Ti 4+ ) to enable simultaneous enrichment and separation of common N-glycopeptides, phosphopeptides, and M6P glycopeptides from tissue/cells. The enrichment was achieved through a dual-mode mechanism based on the electrostatic and hydrophilic properties of the material. The epoxy-ATP-Ti 4+ IMAC material was prepared from epoxy-functionalized silica particles via a convenient two-step process. The ATP molecule provided strong and active phosphate sites for binding phosphopeptides in the conventional IMAC mode and also contributed significantly to the hydrophilicity, which permitted the enrichment of glycopeptides via hydrophilic interaction chromatography. The two modes could be implemented simultaneously, allowing glycopeptides and phosphopeptides to be collected sequentially in a single experiment from the same sample. In addition to standard protein samples, the material was further applied to glycopeptide and phosphopeptide enrichment and characterization from HeLa cell digests and mouse lung tissue samples. In total, 2928 glycopeptides and 3051 phosphopeptides were identified from the mouse lung tissue sample, supporting the utility of this material for large-scale PTM analysis of complex biological samples. Overall, the newly developed epoxy-ATP-Ti 4+ IMAC material and associated fractionation method enable simple and effective enrichment and separation of glycopeptides and phosphopeptides, offering a useful tool to study potential crosstalk between these two important PTMs in biological systems. The MS data have been deposited to the ProteomeXchange Consortium via the PRIDE partner repository with the data set identifier PXD029775.

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Section: Introductionmentioning

confidence: 99%

ATP-Coated Dual-Functionalized Titanium(IV) IMAC Material for Simultaneous Enrichment and Separation of Glycopeptides and Phosphopeptides

Wang,

Huang,

Zhang

et al. 2023

J. Proteome Res.

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“…Support hydrophilicity has been suggested to improve phosphopeptide enrichment selectivity for IMAC 4,10–18 , but also for some MOAC materials 19–22 , metal-organic framework (MOF) materials 23–25 and hybrid MOAC-IMAC materials as well 26 . This observation has prompted the development of hydrophilic materials for phosphopeptide enrichment; for instance, polydopamine was extensively used to prepare MOF 23–25 , IMAC supports 10–14,27–30 , MOAC supports 31 or frameworks further derivatized with linkers, such as adenosine triphosphate 32 . Although some studies were performed by high performance liquid chromatography (HPLC) coupled to mass spectrometry (MS) 11,12,17,23,33 most works in this field analyze samples directly by Matrix-Assisted Laser Desorption/Ionization without any chromatographic separation 10,13,14,16,21,22,24,25,31,32,34,35 , which does not allow a comprehensive characterization of a complex tryptic digest.…”

Section: Introductionmentioning

confidence: 99%

Effect of shell structure of Ti-immobilized metal ion affinity chromatography core-shell magnetic particles for phosphopeptide enrichment

Capriotti

Antonelli

Antonioli

et al. 2019

Sci Rep

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Magnetic materials in sample preparation for shotgun phosphoproteomics offer several advantages over conventional systems, as the enrichment can be achieved directly in solution, but they still suffer from some drawbacks, due to limited stability and selectivity, which is supposed to be affected by the hydrophilicity of the polymeric supports used for cation immobilization. The paper describes the development of an improved magnetic material with increased stability, thanks to a two-step covering of the magnetic core, for the enrichment of phosphopeptides in biological samples. Four materials were prepared featuring a polymeric shell with tunable hydrophilicity, obtained by “grafting from” polymerization of glycidyl methacrylate with 0–8.3% of polyethylene glycol methacrylate (PEGMA), the latter used to modulate the hydrophilicity of the material surface. Finally, the materials were functionalized with iminodiacetic acid for Ti4+ ion immobilization. The materials were analyzed for their composition by a combination of CHN elemental analysis and thermogravimetric analysis, also hyphenated to gas chromatography and mass spectrometric detection. Surface characteristics were evaluated by water contact angle measurements, scanning electron microscopy and energy dispersive X-ray spectrometry. These materials were applied to the enrichment of phosphopeptides from yeast protein digests. Peptides were identified by proteomics techniques using nano-high performance liquid chromatography coupled to mass spectrometry and bioinformatics. Qualitatively the peptides identified by the four systems were comparable, with 1606–1693 phosphopeptide identifications and a selectivity of 47–54% for all materials. The physico-chemical features of the identified peptides were also the same for the four materials. In particular, the grand average of hydropathy index values indicated that the enriched phosphopeptides were hydrophilic (ca. 90%), and only some co-enriched non-phosphorylated peptides were hydrophobic (21–28%), regardless of the material used for enrichment. Peptides had a pI ≤ 7, which indicated a well-known bias for acidic peptides binding, attributed to the interaction with the metal center itself. The results indicated that the enrichment of phosphopeptides and the co-enrichment of non-phosphorylated peptides is mainly driven by interactions with Ti4+ and does not depend on the amount of PEGMA chains in the polymer shell.

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A complementary bimetal synergized with polyethyleneimine functionalized affinity chromatography nanosphere for enrichment of global phosphopeptides

Meng-Yao

Yan

et al. 2022

Chinese Journal of Analytical Chemistry

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A novel IMAC platform – adenosine coupled functional magnetic microspheres for phosphoproteome research

Abstract: Immobilized metal affinity chromatography is a powerful method for phosphopeptide enrichment.

Cited by 4 publications

References 40 publications

ATP-Coated Dual-Functionalized Titanium(IV) IMAC Material for Simultaneous Enrichment and Separation of Glycopeptides and Phosphopeptides

ATP-Coated Dual-Functionalized Titanium(IV) IMAC Material for Simultaneous Enrichment and Separation of Glycopeptides and Phosphopeptides

Effect of shell structure of Ti-immobilized metal ion affinity chromatography core-shell magnetic particles for phosphopeptide enrichment

A complementary bimetal synergized with polyethyleneimine functionalized affinity chromatography nanosphere for enrichment of global phosphopeptides

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