Facile fabrication of Ti⁴⁺-immobilized magnetic nanoparticles by phase-transitioned lysozyme nanofilms for enrichment of phosphopeptides

In this study, titanium (IV)-immobilized magnetic nanoparticles (Ti⁴⁺-PTL-MNPs) were firstly synthesized via a one-step aqueous self-assembly of lysozyme nanofilms for efficient phosphopeptide enrichment. Under physiological conditions, lysozymes readily self-organized into phase-transitioned lysozyme (PTL) nanofilms on Fe₃O₄@SiO₂ and Fe₃O₄@C MNP surfaces with abundant functional groups, including –NH₂, –COOH, –OH, and –SH, which can be used as multiple linkers to efficiently chelate Ti⁴⁺. The obtained Ti⁴⁺-PTL-MNPs possessed high sensitivity of 0.01 fmol μL⁻¹ and remarkable selectivity even at a mass ratio of β-casein to BSA as low as 1:400 for phosphopeptide enrichment. Furthermore, the synthesized Ti⁴⁺-PTL-MNPs can also selectively identify low-abundance phosphopeptides from extremely complicated human serum samples and their rapid separation, good reproducibility, and excellent recovery were also proven. This one-step self-assembly of PTL nanofilms facilitated the facile and efficient surface functionalization of various nanoparticles for proteomes/peptidomes.