Enabling Top-Down Proteomics through Nanotechnology and Materials Chemistry

Proteomics is essential for deciphering how biomolecules interact as a system and for understanding the functions of cellular systems in human diseases. A comprehensive analysis of all proteoforms in the human proteome is essential for gaining a transformative understanding of disease mechanisms and identifying new therapeutic targets. The emerging top-down mass spectrometry (MS)-based proteomics is arguably the most powerful method to comprehensively characterize proteoforms to decipher the post translational modification (PTM) codes together with genetic variations. In this multidisciplinary project in collaboration with Prof. Ying Ge’s group (, we will develop novel approaches enabled by nanotechnology and materials chemistry to address these challenges in top-down MS-based proteomics.

We have developed superparamagnetic Fe3O4 nanoparticles (NPs) whose surface is functionalized by multivalent ligand molecules that specifically bind to the phosphate groups on any phosphoproteins. These NPs enrich phosphoproteins from complex cell and tissue lysates with high specificity. This method enables universal and effective capture, enrichment, and detection of intact phosphoproteins towards a comprehensive analysis of the phosphoproteome using top-down MS. We will further work on developing more multivalent nanoparticles functionalized with ligand groups for specific binding to generally capture and enrich other low-abundance proteins, and will employ these techniques to advance the biological studies using such nanotechnology-enabled top-down proteomics. 

Figure 1:  Synthesis of multivalent nanoparticles functionalized with ligand groups to enrich phosphoproteins for the analysis of the phosphoproteome using top-down MS.


In other directions to address challenges in top-down MS-based proteomics, we have identified ammonium tartrate as a MS-friendly gradient salt, enabling the application of hydrophobic interaction chromatography (HIC), a conventionally MS-incompatible chromatography, to top-down proteomics. We have further developed multi-dimensional liquid chromatography (MDLC) strategies to achieve effective intact protein separation for top-down proteomics. We are currently working on mesoporous silica materials for high-resolution size exclusion chromatography (SEC), and establishing an integrated platform for multi-dimensional intact protein separation strategies with super high resolution.


Figure 2: A novel and effective three dimensional liquid chromatography platform coupling ion exchange chromatography, hydrophobic interaction chromatography and reverse phase chromatography for top-down proteomics.




10) Wenxuan Cai, Trisha Tucholski, Bifan Chen, Andrew J. Alpert, Sean McIlwain, Takushi Kohmoto, Song Jin, and Ying Ge, Top–down Proteomics of Large Proteins up to 223 kDa Enabled by Serial Size Exclusion Chromatography StrategyAnal. Chem.,(2017), DOI: 10.1021/acs.analchem.7b00380

9) Bifan Chen, Leekyoung Hwang, William Ochowicz, Ziqing Lin , Tania Maria Guardado-Alvarez, Wenxuan Cai, Lichen Xiu, Kunal Dani, Cyrus Colah, Song Jin and Ying Ge, Coupling Functionalized Cobalt Ferrite Nanoparticle Enrichment with Online LC/MS/MS for Top-down PhosphoproteomicsChem. Sci.,(2017), DOI: 10.1039/C6SC05435H

8) Leekyoung Hwang, Tania M. Guardado-Alvarez, Serife Ayaz-Gunner, Ying Ge, and Song Jin; A Family of Photolabile Nitroveratryl-Based Surfactants That Self-Assemble into Photodegradable Supramolecular Structures ;Langmuir2016, 32 (16), pp 3963–3969, DOI: 10.1021/acs.langmuir.6b00658

7) Bifan Chen, Ying Peng, Santosh G. Valeja, Lichen Xiu, Andrew J. Alpert, and Ying Ge; Online Hydrophobic Interaction Chromatography–Mass Spectrometry for Top-Down ProteomicsAnal. Chem., 2016, 88 (3), pp 1885–1891, DOI: 10.1021/acs.analchem.5b04285

6) Santosh G. Valeja, Lichen Xiu, Zachery R. Gregorich, Huseyin Guner, Song Jin, and Ying Ge; Three Dimensional Liquid Chromatography Coupling IEC/HIC/RPC for Effective Protein Separation in Top-Down ProteomicsAnal. Chem.2015, 87 (10), pp 5363–5371, DOI: 10.1021/acs.analchem.5b00657

5). Ying-Hua Chang, Zachery r. Gregorich, Albert J. chen, Leekyoung Hwang, Huseyin Guner, Dyang Yu, Jianyi, and Ying Ge; New Mass-Spectrometry-Compatible Degradable surfactant for Tissue ProteomicsJournal of Proteome Res. 2015, 14 (3), pp 1587–1599 DOI: 10.1021/pr5012679

4). Leekyoung Hwang, Serife Ayaz-Guner, Zachery R. Gregorich, Wenxuan Cai, Santosh G. Valeja, Song Jin, and Ying Ge;  Specific Enrichment of Phosphoproteins Using Functionalized Multivalent NanoparticlesJ. Am. Chem. Soc.2015, 137 (7), 2432-2435, DOI: 10.1021/ja511833y

3). Lichen Xiu, Santosh G. Valega, Andrew J. Alpert, Song Jin, and Ying Ge; Effective Protein Separation by Coupling Hydrophobic Interaction and Reverse Phase Chromatography for Top-down ProteomicsAnal. Chem.2014, 86 (15), pp 7899-7906, DOI: 10.1021/ac501836k

2). Nelson, C. A.; Szezech, J. R.; Zhu, H.; Xu, Q.; Lawrence, M. J.; Jin, S.; Ge, Y.; Effective Enrichment and Mass Spectrometry Analysis of Phosphopeptides Using Mesoporous Metal Oxide Nanomaterials, Anal. Chem. 2010, 82, 7193-7201.

1). Nelson, C. A.; Szczech, J. R.; Xu, Q.; Lawrence, M. J., Jin, S.; Ge, Y.; Mesoporous Metal Oxide Nanomaterials Effectively Enrich Phosphopeptides for Mass Spectrometry-based Phosphoproteomics, Chem. Commun. (2009) 6607-6609


1). Song Jin; Ying Ge; Cory Nelson, Qingge Xu; Use of nanomaterials to Enrich Phosphopeptides for Mass Spectrometry-Based Proteomics, 2013