A proteome is a complete set of proteins synthesized in an organism, tissue, or cells. It is often referred to as the proteome of a species (e.g., Homo sapiens) or organs (e.g., kidneys). Proteomics deals with the study of proteomes.
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One of the most common analytical techniques used in proteomics is mass spectrometry (MS). The application of nanoparticles (NPs) in proteome analysis has enabled targeted and non-targeted proteome analysis.
What is Proteomic Analysis?
Proteomics is involved with investigating the proteome composition and its function in an organism. Proteins play the most important role in cellular activities and are regarded as critical macromolecules in precision medicine.
The advancements in MS-based proteomics concerning its sensitivity, accuracy, automation, and throughput have significantly contributed to the discovery of novel biomarkers for many diseases. Additionally, proteomic analysis has shed light on the underlying molecular mechanisms of many diseases.
Several challenges have surfaced during the global characterization of protein expression using targeted proteomics, particularly in examining the spatial and temporal organization of the proteome. It is difficult to analyze the proteome because of its dynamic nature and varied complexities. Furthermore, the heterogeneity in protein abundance is condition-dependent and subject to post-translational modifications (PTMs).
Application of Nanomaterials in Proteomic Study
Nanoparticles (NPs) are extremely small particles whose size ranges between 1 and 100-nanometer range. Some of the unique properties of NPs, such as good biocompatibility, large specific surface area, abundant active affinity sites, and ultra-small size, make them appropriate for proteomic applications.
To reduce sample complexity and enable deep proteome coverage, it is important to pre-treat the samples before MS analysis. Before analysis, proteins of interest must be isolated from the complex mixture. For example, magnetic nanocomposites have been effectively applied for protein purification, which significantly decreases the processing time. Mesoporous nanoparticles are also used for size-based separation and membrane proteomics.
Antibiotic-based enrichment methods are used to determine protein acetylation, which plays a vital role in regulating chromatin accessibility, gene transcription protein localization, and enzymatic activity. This method substantially increases the chance of detecting important proteins available in low concentrations.
Nanoparticles can selectively or non-selectively bind to proteins during global proteome profiling. For example, iron and titanium oxide nanoparticles are used to identify targeted proteins with specific PTMs (e.g., glycosylation and phosphorylation), while titanium dioxide nanorods are used for non-selective analysis.
Nanotechnology-based Targeted Proteome Analysis
As stated above, two widely studied PTMs are phosphorylation and glycosylation. Phosphorylation regulates cell signaling and plays a key role in the disease-causing irregularities of kinase signaling networks. Due to low stoichiometry, proteomic analysis of phosphorylation is not an easy task.
Two widely used techniques, metal oxide affinity chromatography (MOAC) and immobilized metal ion affinity chromatography (IMAC), used for phosphoproteomic studies are associated with magnetic nanoparticles. IMAC technique uses various metallic nanoparticles, including aluminum, iron, titanium, and gallium.
Glycosylation is another structurally complex protein modification process. Based on linkage sites, protein glycosylation is divided into two types, i.e., N-glycosylation and O-glycosylation. A large body of research indicated that cancer cells have distinct glycosylation profiles. A modified glycosylation profile serves as a hallmark of cancer.
Both affinity-based and covalent binding-based enrichment methods are glycosylation enrichment techniques that utilize nanomaterials. The affinity-based methods deal with the immobilization of lectin onto the surface of NPs. In the covalent binding-based method, glycopeptides are typically enriched by the interaction between diol-containing glycoproteins/glycopeptide and boronic acid.
Cysteine is inclined to post translational modifications because of its high reactivity and redox sensitivity. Some common PTMs on cysteine are lipid-derived electrophilic 4-hydroxynonenal, and redox-dependent nitrosylation (SNO). A novel strategy based on nanographite fluoride has been developed to study SNO-peptides.
Nanotechnology-Based Non-Targeted Proteome Analysis
Nanoparticle-based non-specific enrichment methods present important alternatives to conventional targeted proteomic analysis.
Protein corona is a group of proteins that are instantaneously and non-specifically adsorbed onto the nanoparticle surface when exposed to biospecimens. Protein corona formation alters the physicochemical properties of nanoparticles.
Protein corona is not commonly used in the discovery of important biomarkers. A recent protein corona profiling revealed that it represented a subset of the plasma proteome. Since the composition of protein corona varies in accordance with nanoparticle properties, scientists assume that engineered NPs can capture distinct protein corona patterns for plasma proteome profiling. Several types of magnetic nanoparticles can perform in-depth plasma proteome profiling.
Circulating tumor cells (CTCs) are released from primary or metastatic tumors into the circulation system. Detection of CTCs offers a unique opportunity to identify markers associated with various cancers and their stages. Non-magnetic beads (e.g., titanium dioxide nanorods) are used to examine CTCs.
Utilization of NPs in Blood Proteomics
Blood is the most common body fluid, which hosts a wide arrange of molecules, including small molecules, electrolytes, drugs, and proteins that are routinely tested in clinical settings. MS-based proteomics is extremely challenging and is used to detect novel blood biomarkers without fractionation, degradation, or enrichment. Liquid Chromatography-MS/MS can only identify a few hundred proteins.
Nanoparticles have been widely applied in blood proteomics. A classical antibody-immobilized magnetic nanomaterials-based method, namely, Stable Isotope Standards and Capture by Anti-Peptide Antibodies (SISCAPA), is used to capture specific blood protein biomarkers.
The development of targeted proteomics methods, such as multiple reaction monitoring (MRM), has reduced the probability of false negative results in the early diagnosis of hepatocellular carcinoma. Application of surface-functionalized superparamagnetic iron-oxide (magnetite) can selectively enrich cardiac biomarker troponin I (cTnI).
Compared to conventional methods, nanoparticle-based methods with unique characteristics, such as superior biocompatibility, large specific surface area, abundant active affinity sites, and unique surface properties, are favorable for proteomic analysis. As the majority of enrichment methods are carried out in vitro, in the future, the development of in vivo enrich technology is required. This technology must be equipped to provide the spatial location and functional information about the protein under consideration. This information will aid in the creation of precision medicine.
References and Further Reading
Zhang, Y. et al. (2022). Application of nanomaterials in proteomics-driven precision medicine. Theranostics, 12(6), pp. 2674–2686. https://doi.org/10.7150/thno.64325
Ferdosi, S. et al. (2022) Engineered nanoparticles enable deep proteomics studies at scale by leveraging tunable nano–bio interactions. Applied Biological Sciences, 119(11), p. e2106053119. https://doi.org/10.1073/pnas.2106053119
Blume, J.E. et al. (2020) Rapid, deep and precise profiling of the plasma proteome with multi-nanoparticle protein corona. Nature Communications, 11, p. 3662. https://doi.org/10.1038/s41467-020-17033-7
Vogt, C. et al. (2015) Proteomics Analysis Reveals Distinct Corona Composition on Magnetic Nanoparticles with Different Surface Coatings: Implications for Interactions with Primary Human Macrophages. PLoS ONE, 10(10), p. e0129008. https://doi.org/10.1371/journal.pone.0129008
Marko,F. N. et al. (2007) Nanotechnology in proteomics. Expert Review of Proteomics, 4(5), pp. 617-626, https://www.tandfonline.com/doi/full/10.1586/147894220.127.116.117
Geho, H. D. et al. (2007) Nanotechnology in clinical proteomics. Future Medicine, 2(1), pp. 1-5. https://doi.org/10.2217/17435818.104.22.168