Quantitation of the Noncovalent Cellular Retinol-Binding Protein, Type 1 Complex Through Native Mass Spectrometry
JournalJournal of the American Society for Mass Spectrometry
PublisherSpringer New York LLC
MetadataShow full item record
AbstractNative mass spectrometry (MS) has become a valuable tool in probing noncovalent protein–ligand interactions in a sample-efficient way, yet the quantitative application potential of native MS has not been fully explored. Cellular retinol binding protein, type I (CrbpI) chaperones retinol and retinal in the cell, protecting them from nonspecific oxidation and delivering them to biosynthesis enzymes where the bound (holo-) and unbound (apo-) forms of CrbpI exert distinct biological functions. Using nanoelectrospray, we developed a native MS assay for probing apo- and holo-CrbpI abundance to facilitate exploring their biological functions in retinoid metabolism and signaling. The methods were developed on two platforms, an Orbitrap-based Thermo Exactive and a Q-IMS-TOF-based Waters Synapt G2S, where similar ion behaviors under optimized conditions were observed. Overall, our results suggested that within the working range (~1–10 μM), gas-phase ions in the native state linearly correspond to solution concentration and relative ion intensities of the apo- and holo-protein ions can linearly respond to the solution ratios, suggesting native MS is a viable tool for relative quantitation in this system.
SponsorsThis project was funded with federal funds from NIH: R01HD077260 and NIH/NIAID contract no. HHSN272201000046C. Additional support was provided by the University of Maryland, School of Pharmacy, Mass Spectrometry Center (SOP1841-IQB2014).
KeywordCellular retinol-binding protein
Native mass spectrometry
Noncovalent protein complex
Identifier to cite or link to this itemhttps://www.scopus.com/inward/record.uri?eid=2-s2.0-85006355611&doi=10.1007%2fs13361-016-1499-5&partnerID=40&md5=dc6a309c473acefbc1f321841fe1594e; http://hdl.handle.net/10713/11074