Nianrong Sun

Size-exclusive magnetic graphene/mesoporous silica composites with titanium(IV)-immobilized pore walls for selective enrichment of endogenous phosphorylated peptides.

Developing an effective separation method is necessary for identifying low-abundant endogenous phosphorylated peptides with the removal of proteins. In this work, we prepared size-exclusive magnetic graphene/mesoporous silica composites with titanium(IV)-immobilized pore walls (denoted as Ti(4+)-MGMSs) for capturing endogenous phosphorylated peptides for mass spectrometry analysis. The introduction of hydrophilic polydopamine simplified the synthetic process of Ti(4+)-MGMSs, and the ordered mesoporous channels are beneficial to the trapping of endogenous phosphopeptides while large-size proteins are excluded. Furthermore, the magnetic performance greatly simplifies the entire process of enrichment. With all of the advances, the novel Ti(4+)-MGMSs present high enrichment efficiency either from the low concentration of β-casein tryptic digest (0.5 fmol/μL) or the mixture of β-casein tryptic digest and α-casein (or plus bovine serum albumin, with a mass ratio up to 1:500). Besides, Ti(4+)-MGMSs have also been successfully applied to enrich endogenous phosphorylated peptides from human serum and human saliva.

Highly selective enrichment of N-linked glycan by carbon-functionalized ordered graphene/mesoporous silica composites.

Abnormal protein glycosylation has been demonstrated to be associated with many diseases; therefore, it is very important to conduct a comprehensive structure analysis of glycan for prognosis and diagnosis of diseases, such as cancer. In this work, for the first time, carbon-functionalized ordered graphene/mesoporous silica composites (denoted as C-graphene@mSiO2) with large surface area and uniform pore size were designed and synthesized. By taking advantage of the special interaction between the carbon and glycans as well as size-exclusion ability, 25 N-linked glycans released from ovalbumin were observed clearly with strong MS signals and increased signal-to-noise (S/N) ratio. In addition, after enrichment with the C-graphene@mSiO2 composites, 48 N-linked glycans (S/N > 10) with sufficient peak intensities were obtained from only 400 nL of healthy pristine human serum. The facile and low-cost synthesis method as well as high selective enrichment ability of the novel C-graphene@mSiO2 composite makes it a promising tool for glycosylation research.

Hydrophilic Mesoporous Silica Materials for Highly Specific Enrichment of N-Linked Glycopeptide

Hydrophilic interaction liquid chromatography (HILIC) is a significant enrichment strategy in glycoproteomics profiling. In this report, hydrophilic magnetic mesoporous silica materials (denoted as Fe3O4@mSiO2-IDA) were designed and synthesized as an outstanding enrichment platform for glycopeptide analysis. By taking advantage of their merits, such as large surface area, excellent hydrophilicity and unbiased affinity toward all types of glycopeptides, the Fe3O4@mSiO2-IDA nanomaterials were successfully applied to capture glycopeptides from complex samples. A total of 25 glycopeptides from horseradish peroxidase (HRP) digests and 33 glycopeptides from IgG were identified, respectively. Especially, as a result, 424 glycopeptides assigned to 140 glycoproteins were identified from only 2 μL human serum.

Rapid synthesis of titanium(IV)-immobilized magnetic mesoporous silica nanoparticles for endogenous phosphopeptides enrichment

The MALDI‐TOF MS has already been a main platform for phosphoproteome analysis. However, there are some weaknesses in direct analysis of endogenous phosphopeptides by MALDI‐TOF MS because of the serious suppression effect and poor ionization efficiency, which is brought by the excess of nonphosphopeptides and protein. It is essential to enrich endogenous phosphopeptides from complex biosamples efficiently prior to MALDI‐TOF MS analysis. Herein, we present a time‐saving and detailed protocol for the synthesis of titanium(iv)‐immobilized magnetic mesoporous silica nanoparticles (denoted as Fe3O4@mSiO2‐Ti4+), the subsequent enrichment process, and MALDI‐TOF MS analysis. We tested the LOD, size‐exclusive effect, reproducibility, and stability of Fe3O4@mSiO2‐Ti4+ nanoparticles. Furthermore, the ability of this protocol for identifying endogenous phosphopeptides in healthy human serum and saliva was investigated.

Facile synthesis of thiol-polyethylene glycol functionalized magnetic titania nanomaterials for highly efficient enrichment of N-linked glycopeptides

As protein N-glycosylation involved in generation and development of various cancers and diseases, it is vital to capture glycopeptides from complex biological samples for biomarker discovery. In this work, by taking advantages of the interaction between titania and thiol groups, thiol-polyethylene glycol functionalized magnetic titania nanomaterials (denoted as Fe3O4@TiO2@PEG) were firstly fabricated as an excellent hydrophilic adsorbent of N-linked glycopeptides. On one hand, the special interaction of titanium-thiol makes the synthetic manipulation simple and provides a new idea for design and synthesis of novel nanomaterials; on the other hand, strong magnetic response could realize rapid separation and the outstanding hydrophilicity of polyethylene glycol makes Fe3O4@TiO2@PEG nanomaterials show superior performance for glycopeptides enrichment with ultralow limit of detection (0.1mol/μL) and high selectivity (1:100). As a result, 24 and 33 glycopeptides enriched from HRP and IgG digests were identified respectively by MALDI-TOF MS, and 300 glycopeptides corresponding to 106 glycoproteins were recognized from merely 2μL human serum, indicating a great potential of Fe3O4@TiO2@PEG nanomaterials for glycoproteomic research.

Glucose-6-Phosphate-Functionalized Magnetic Microsphere as Novel Hydrophilic Probe for Specific Capture of N-Linked Glycopeptides

Developing cost-effective approaches based on hydrophilic interaction liquid chromatography (HILIC) has been the main tendency for low-abundance glycopeptides capture before LC-MS/MS analysis. Carbohydrates with outstanding biocompatibility and hydrophilicity are ubiquitous in the kingdoms of animal and plant and could be a wonderful choice as functional groups for glycopeptides enrichment. In this work, glucose-6-phosphate, as one of the indispensable cogs in pivotal metabolic wheels of life, was chosen as functionalized groups to be grafted onto the surface of Fe3O4 microspheres via one-step surface fabrication strategy. The acquired hydrophilic Fe3O4@G6P microspheres showed superior enrichment performance for glycopeptides with high sensitivity (0.5 fmol/μL) and high selectivity (1:100) and good repeatability (10 times at least). Furthermore, the Fe3O4@G6P microspheres also exhibited enrichment ability for glycopeptides in different biosamples. A total of 243 glycopeptides assigned to 92 glycoproteins and 183 glycopeptides corresponding to 74 different glycoproteins was identified from merely 2 μL of serum and saliva, respectively.

Hydrophilic Probe in Mesoporous Pore for Selective Enrichment of Endogenous Glycopeptides in Biological Samples

As one of the most important post-translational modifications (PTMs) of peptidome, glycopeptidome is closely related to serious disease, especially to cancer. In order to specifically discover and analyze glycopeptidome biomarkers for clinical diagnosis of cancer on early-stage, it is crucial to develop efficient technique to analyze low-abundance of endogenous glycopeptides in biological samples. In this report, a hydrophilic probe in mesoporous pore (denoted as Fe3O4@mSiO2@G6P) was designed and prepared. By taking advantage of the excellent hydrophilicity and size-exclusion ability, we applied Fe3O4@mSiO2@G6P to capture glycopeptides from both horseradish peroxidase (HRP) and immunoglobulin (IgG) digests successfully. Moreover, a total of 39 and 25 endogenous glycopeptides were identified from healthy saliva and gastric saliva, respectively, indicating the great potential of this probe for the exploration of glycopeptidome biomarkers.