title: Application of ion mobility-mass spectrometry to microRNA analysis.
abstract: Liquid chromatography/mass spectrometry is widely used for studying sequence determination and modification analysis of small RNAs. However, the efficiency of liquid chromatography-based separation of intact small RNA species is insufficient, since the physiochemical properties among small RNAs are very similar. In this study, we focused on ion mobility-mass spectrometry (IM-MS), which is a gas-phase separation technique coupled with mass spectrometry; we have evaluated the utility of IM-MS for microRNA (miRNA) analysis. A multiply charged deprotonated ion derived from an 18-24-nt-long miRNA was formed by electrospray ionization, and then the time, called the "drift time", taken by each ion to migrate through a buffer gas was measured. Each multivalent ion was temporally separated on the basis of the charge state and structural formation; 3 types of unique mass-mobility correlation patterns (i.e., chainlike-form, hairpin-form, and dimer-form) were present on the two-dimensional mobility-mass spectrum. Moreover, we found that the ion size (sequence length) and the secondary structures of the small RNAs strongly contributed to the IM-MS-based separation, although solvent conditions such as pH had no effect. Therefore, sequence isomers could also be discerned by the selection of each specific charged ion, i.e., the 6(-) charged ion reflected a majority among chainlike-, hairpin-, and other structures. We concluded that the IM-MS provides additional capability for separation; thus, this analytical method will be a powerful tool for comprehensive small RNA analysis.
authors.affiliation: Department of Biotechnology, Graduate School of Engineering, Osaka University, 2-1 Yamadaoka, Suita, Osaka, Japan.
authors.forename: Kosuke
authors.initials: K
authors.lastname: Takebayashi
authors.forename: Kenji
authors.initials: K
authors.lastname: Hirose
authors.forename: Yoshihiro
authors.initials: Y
authors.lastname: Izumi
authors.forename: Takeshi
authors.initials: T
authors.lastname: Bamba
authors.forename: Eiichiro
authors.initials: E
authors.lastname: Fukusaki
mesh_terms: D001483:Base Sequence; D007477:Ions; D007536:Isomerism; D035683:MicroRNAs / Q000737:chemistry / Q000302:isolation & purification*; D009690:Nucleic Acid Conformation; D012997:Solvents; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*
chemical_list: D007477:Ions; D035683:MicroRNAs; D012997:Solvents