Self‐Assembled TiO₂ Nanocrystal Clusters for Selective Enrichment of Intact Phosphorylated Proteins

Abstract

Mesoporous oxides have been used for selective separation and adsorption of biomolecules. [1][2][3][4][5][6][7][8][9][10][11] Inorganic oxide materials such as silica can be synthesized into highly ordered mesoporous structures featuring high in-pore surface areas, narrow pore size distributions, adjustable pore sizes, and modifiable surface properties. [10,[12][13][14][15] Such unique properties favor the selective enrichment of proteins or peptides based on the size-exclusion mechanism. Classic mesoporous silica structures with narrow pore size distributions, such as MCM-41 and SBA-15, have been used to selectively enrich peptides from human plasma and exclude other proteins according to an accurate molecular weight (MW) cutoff. [2][3][4][5][6][7][8][9] However, this type of enrichment method is limited mainly to silica and alumina because currently it is still difficult to extend broadly the surfactant-templating method to the synthesis of mesoporous structures of many other oxides in a convenient, controllable, and scalable manner (although the literature does contain some relevant procedures [16][17][18] ). The limited choice of materials makes it difficult to further enhance the selectivity by taking advantage of both the size-exclusion effect and the specific binding between many oxides and proteins or peptides.Herein, we propose a general strategy for the fabrication of novel porous nanostructured materials for the efficient separation of biomolecules such as proteins, peptides, and DNA. Briefly, nanoparticles of various nanostructured materials with uniform sizes and shapes are firstly synthesized and then self-assembled into three-dimensional submicrometer clusters containing uniform mesoscale pores. Thanks to rapid progress in colloidal nanostructure synthesis, a great number of materials can now be routinely produced in the form of nanoparticles with excellent control over size, shape, and surface properties, [19][20][21][22] making it possible to utilize specific material-protein/peptide interactions to enhance the selectivity in protein/peptide enrichment. [23,24] Additionally, size exclusion can also be achieved by controlling the dimensions of the pores produced by the packing of constituent nanoparticles, [25,26] allowing selective enrichment of biomolecules based on their sizes. The outer surface of each cluster can be made highly hydrophilic, so that nonspecific binding of many hydrophobic proteins/peptides can be avoided. The selfassembly process also brings the convenience of incorporation of multiple components into the clusters to further facilitate separation and detection. For example, the incorporation of fluorescent nanocrystals, such as quantum dots, during the assembly process may produce multifunctional microspheres that are able not only to selectively enrich but also to easily identify the targeted biomolecules. Also, adding superparamagnetic iron oxide nanocrystals to the clusters allows their efficient removal from the analyte solution after selective adsorption by using an exter...

Self-assembly and photocatalysis of mesoporous TiO2 nanocrystal clusters

“…During the initial adsorption process, ~30% of RhB was adsorbed by the silicatreated clusters, while only 20% of RhB could be adsorbed by the untreated clusters. We have pointed out previously that the silica coating and etching processes enhances the surface charge and water dispersity of such clusters [40], which is confirmed by these experiments. Without the silica coating/removing treatment, the calcined TiO 2 clusters are in the form of large aggregates and cannot be well dispersed in water even after 10 min of sonication, while the silicatreated clusters can be well dispersed in water.…”

“…Recently, we and other groups reported a general strategy for the fabrication of novel porous nanostructured materials based on the self-assembly of nanocrystals [38][39][40]. We have demonstrated that the as-prepared mesoporous nanocrystal clusters can be used for selective enrichment of peptides and proteins from complex biological samples [40][41][42].…”

“…We have demonstrated that the as-prepared mesoporous nanocrystal clusters can be used for selective enrichment of peptides and proteins from complex biological samples [40][41][42]. From the perspective of photocatalysis, there are also several attractive features of these materials, including clean surfaces, high crystallinity, small grain size, and large surface area even after high temperature treatment, good water dispersity, and a pore size distribution which is controllable by simply tuning the initial nanocrystal size and shape.…”

Recent advances of mesoporous materials in sample preparation

“…10. Illustration of (a) the procedure for the fabrication of the mesoporous TiO2 nanocrystal clusters and (b) the selective enrichment process of phosphorylated proteins using the mesoporous TiO2 clusters and the elutes were analyzed by CE or MALDI-TOF MS [97].…”

“…10, Lu et al [97] described the preparation self-assembled TiO 2 nanocrystal clusters with different averaged pore sizes of 2.3, 2.5 and 3.5 nm, and their application on selective enrichment of phosphorylated proteins from a mixed solution of three proteins (one phosphorylated ␤-casein, two non-phosphorylated horseradish peroxidase and ␤-lactoglobin). Only ␤-casein was detected by capillary electrophoresis with UV detection at 200 nm when using the mesoporous TiO 2 clusters.…”