Ti⁴⁺‐Immobilized Magnetic Composite Microspheres for Highly Selective Enrichment of Phosphopeptides

Abstract

Architectural design is essential to achieve ideal chemical and biological properties of nanomaterials. In this article, a novel route to fabricate high‐quality magnetic composite microspheres composed of a high‐magnetic‐response magnetic colloid nanocrystal cluster (MCNC) core, a poly(methylacrylic acid) (PMAA) interim layer, and a Ti4+‐immobilized poly(ethylene glycol methacrylate phosphate) (PEGMP) shell via two‐step distillation–precipitation polymerization is presented. The unique as‐synthesized MCNC@PMAA@PEGMP‐Ti4+ composite microsphere is investigated for its applicability for selective enrichment of phosphopeptides from complex biological samples. The experiment results demonstrate that, by taking advantage of the pure phosphate–Ti4+ interface and high Ti4+ loading amount, the MCNC@PMAA@PEGMP‐Ti4+ composite microsphere possesses remarkable selectivity for phosphopeptides even at a very low molar ratio of phosphopeptides/nonphosphopeptides (1:500). The extreme sensitivity, excellent recovery of phosphopeptides, and high magnetic susceptibility are also proven. These outstanding features demonstrate that the MCNC@PMAA@PEGMP‐Ti4+ composite microspheres have great benefit for the pretreatment before mass spectrometric analysis of phosphopeptides. Furthermore, the performance of the approach in selective enrichment of phosphopeptides from drinking milk and human serum gives powerful evidence for its high selectivity and effectiveness in identifying the low‐abundant phosphopeptides from complicated biological samples.

Preparation of on-plate immobilized metal ion affinity chromatography platform via dopamine chemistry for highly selective isolation of phosphopeptides with matrix assisted laser desorption/ionization mass spectrometry analysis

“…In order to demonstrate the feasibility of our method to complex samples, we carried out the onplate enrichment of phosphopeptide from human serum, which is expected containing four endogenous phosphopeptides with molecular masses of 1389, 1460, 1545 and 1616 Da coming from fibrinogen α-chain [28][29][30][31]. Because a large amount of nonphosphopeptides, abundant proteins and inorganic salts existing in the serum increases the complexity of condition largely, the detection of endogenous phosphopeptides contained in human serum will be suppressed, which results in almost no peptides found in the spectrum before enrichment (Fig.…”

Aluminium glycinate functionalized silica nanoparticles for highly specific separation of phosphoproteins

“…[5][6][7][8][9][10][11][12] Nevertheless, this bottom-up proteomics also suffers from drawbacks such as loss of labile PTMs and sequence variants. 13 In contrast, top-down MS analysis of intact phosphoproteins could provide a better description of protein phosphorylation 14,15 and regain considerable attention with new MS instruments such as Fourier transform ion cyclotron resonance 16,17 and Orbitrap MS 18,19 .…”

Highly specific purification of N-glycans using phosphate-based derivatization as an affinity tag in combination with Ti4+-SPE enrichment for mass spectrometric analysis

“…The Ti 4þ -immobilized magnetic composite microspheres were homemade, detailed synthesis route and characterizations can be referred to our previous publication [16]. Briefly, a high-magneticresponse magnetic colloid nanocrystal cluster (MCNC) core with the size of approximately 300 nm were prepared, and then a poly(methylacrylic acid) (PMAA) interim layer were covered on the MCNC core by distillation-precipitation polymerization.…”