Wisconsin Alumni Research Foundation

Ying Ge

Song Jin

Kyle Brown

Tania Guardado Alvarez

Cory Alexander Nelson

Qingge Xu

Kevin Buck

The present invention provides photo-cleavable anionic surfactants, particularly 4-hexylphenylazosulfonate (Azo) and sodium 4-hexylphenylazosulfonate derivatives, which can be rapidly degraded upon UV irradiation, for top-down and bottom-up proteomics. These surfactants can effectively solubilize proteins and peptide fragments with performance comparable to sodium dodecyl sulfate (SDS) and are compatible with mass spectrometry analysis of the solubilized proteins and peptide fragments. Top-down proteomic studies using the present photo-cleavable anionic surfactants has allowed the detection of 100-fold more unique proteoforms as compared to controls and has enabled the solubilization of membrane proteins for comprehensive characterization of protein post-translational modifications. In addition, the present photo-cleavable anionic surfactants are also suitable for dissolving polypeptides in bottom-up proteomic experiments including extracellular matrix proteomics, and are suitable as a substitute for SDS in gel electrophoresis.

Photo-cleavable surfactants

Exosomes are a small type of extracellular vesicles containing nucleic acids, lipids, and proteins that are implicated in tumorigenesis, metastasis, and cardiac regeneration, and therefore serve as potentially useful biomarkers from fluids or as vehicles for drug delivery. Global bottom-up mass spectrometry-based proteomics has been previously used to profile exosome cargo for diagnostic purposes. However, the current protocols for MS analysis of extracellular vesicles and exosome proteomics are challenging due to labor-intensive sample preparation, including lengthy digestion times and removal of MS incompatible reagents, and the need for high sensitivity. To address these challenges, the present invention provides a novel, high-throughput strategy for extracellular vesicle analysis and exosome proteomics using a photo-cleavable, anionic surfactant, preferably 4-hexylphenylazosulfonate (Azo). These photo-cleavable, anionic surfactants are able to facilitate high-throughput digestion with minimal sample cleanup for quantitative and highly reproducible analysis.

Strategy enabled by a photo-cleavable surfactant for extracellular vesicle proteomics

The present invention provides methods and materials for isolating, purifying, and/or enriching the concentration of compounds having one or more phosphate groups and/or derivatives thereof, including but not limited to phosphorylated peptides and/or phosphorylated proteins. In some aspects, the present invention provides nanostructured enrichment materials, such as metal oxide mesoporous materials, that selectively and reversibly bind with phosphorylated compounds with high specificity and are capable of controlled release of phosphorylated compounds bound to their active surfaces. Mesoporous materials of the present invention also provide enrichment materials having large active surface areas that provide for higher loading capacities for phosphorylated peptides and proteins relative to conventional affinity based methods. Nanostructured metal oxide mesoporous enrichment materials of the present invention are also compatible with implementation via a variety of separation platforms including flow through separation systems, elution based separation systems, column chromatography and affinity chromatography.

Growing community of inventors

Madison, WI, United States of America

Ying Ge