title: Polarity-based fractionation in proteomics: hydrophilic interaction vs reversed-phase liquid chromatography.
abstract: During recent decades, hydrophilic interaction liquid chromatography (HILIC) ahs been introduced to fractionate or purify especially polar solutes such as peptides and proteins while reversed-phase liquid chromatography (RPLC) is also a common strategy. RPLC is also a common dimension in multidimensional chromatography. In this study, the potential of HILIC vs RPLC chromatography was compared for proteome mapping of human peripheral blood mononuclear cell extract. In HILIC a silica-based stationary phase and for RPLC a C18 column were applied. Then separated proteins were eluted to an ion trap mass spectrometry system. Our results showed that the HILIC leads to more proteins being identified in comparison to RPLC. Among the total 181 identified proteins, 56 and 38 proteins were fractionated specifically by HILIC and RPLC, respectively. In order to demonstrate this, the physicochemical properties of identified proteins such as polarity and hydrophobicity were considered. This analysis indicated that polarity may play a major role in the HILIC separation of proteins vs RPLC. Using gene ontology enrichment analysis, it was also observed that differences in physicochemical properties conform to the cellular compartment and biological features. Finally, this study highlighted the potential of HILIC and the great orthogonality of RPLC in gel-free proteomic studies. Copyright © 2015 John Wiley & Sons, Ltd.
authors.affiliation: Drug Design and Bioinformatics Unit, Medical Biotechnology Department, Biotechnology Research Center, Pasteur Institute of Iran, Tehran, Iran.
authors.forename: M
authors.initials: M
authors.lastname: Jafari
authors.affiliation: Department of Computational Biology, Faculty of High Technologies, Tarbiat Modares University, Tehran, Iran.
authors.forename: M
authors.initials: M
authors.lastname: Mirzaie
authors.affiliation: National Institute of Genetic Engineering and Biotechnology, Tehran, Iran.
authors.forename: M
authors.initials: M
authors.lastname: Khodabandeh
authors.affiliation: Department of Phytochemistry, Medicinal Plants and Drugs Research Institute, Shahid Beheshti University, Tehran, Iran.
authors.forename: H
authors.initials: H
authors.lastname: Rezadoost
authors.affiliation: Department of Phytochemistry, Medicinal Plants and Drugs Research Institute, Shahid Beheshti University, Tehran, Iran.
authors.forename: A
authors.initials: A
authors.lastname: Ghassempour
authors.affiliation: Pharmaceutical and Medicinal Chemistry Department, Pharmaceutical and Drug Industries Research Division, National Research Centre, Dokki, 12622, Giza, Egypt.
authors.forename: H Y
authors.initials: HY
authors.lastname: Aboul-Enein
mesh_terms: D056148:Chromatography, Reverse-Phase / Q000379:methods*; D006801:Humans; D057927:Hydrophobic and Hydrophilic Interactions; D040901:Proteomics
keywords: hydrophilic interaction liquid chromatography; hydrophobicity; orthogonality; polarity; proteome mapping