Highly Selective Capture of Monophosphopeptides by Two-Dimensional Metal-Organic Framework Nanosheets

Separation of monophosphopeptides from multi-phosphopeptides in complex biological samples is significant in the study of protein kinase signal transduction pathways. To the best of our knowledge, very few materials have been reported that could selectively enrich monophosphopeptides because of the chemical difficulty in retaining the intermediate monophosphopeptides and excluding both non-phosphopeptides and multi-phosphopeptides in acidic conditions, which requires unique interactions to balance the metallic affinity and the hydrophobicity. With the large surface area, abundant accessible active sites, and ultrathin structures, two-dimensional (2-D) metal-organic framework (MOF) Hf-1,3,5-tris(4-carboxyphenyl)benzene (BTB) nanosheets were rationally selected. Due to the elongated organic ligands and the balance between metallic affinity of clusters and hydrophobicity from ligands, the 2-D Hf-BTB nanosheets exhibited unique enrichment selectivity toward monophosphopeptides. The 2-D MOF nanosheets demonstrated excellent sensitivity (detection limit of 0.4 fmol μL^-1) and selectivity [1:1000 molar ratios of β-casein/BSA (bovine serum albumin)] in model phosphopeptides enrichment. The nanosheets were implemented for the analysis of nonfat milk and human saliva samples as well as in situ isotope labeling for dysregulated phosphopeptides from patients' serum with anal canal inflammation, exhibiting 6.6-fold upregulation of serum phosphopeptide HS4 (ADpSGEGDFLAEGGGVR) compared to the control healthy serum. The proteomics analysis of mouse brain cortical samples associated with Alzheimer's disease, which were from Akt (protein kinase B) conditional knockout mouse and littermate control mouse, was further established with 2-D Hf-BTB nanosheets. With high capture efficiency for monophosphopeptides, this method was capable of distinguishing the difference of monophosphopeptides from microtubule-associated protein τ (MAPT/τ) between the Akt knockout sample and control sample.