AbstractThe technique of ion mobility mass spectrometry (IM-MS) measures the mass-to-charge ratio of a sample and also allows the mobilities of ions in inert drift gases to be measured prior to conversion to their collision cross sections (CCS). The CCS of a molecule relates to its three-dimensional size and shape. Some ion mobility spectrometers require calibration with appropriate standards to obtain accurate CCS values from measured mobilites in the gas of choice. Many calibration systems do not encompass the full range of mass and CCS values of interest. Polymer Factory have developed a discrete set of standards that increases the possible calibration range, known as SpheriCalÂ®. Consequently, SpheriCalÂ® allows accurate calibration for a greater range of potential analytes than calibrant systems that are currently commercially available. Although most traditional calibration systems (polyalanine, ultramark, LCMS QC reference, and tetraalkyl ammonium salts) can calibrate below CCS values of 400 Ã 2 and 2000 m/z, the main aim of SpheriCalÂ® is to provide values as high as 800 Ã 2 and 5500 m/z. This will greatly increase the range from which accurate CCS data may be obtained. Ion mobility data was acquired on two commercially available instruments namely an Agilent 6560 (IM-QToF) and SYNAPT G2 HDMS (DTIMS) in both cases using nitrogen as the drift gas. Data was acquired as drift times which were then analysed using MassHunter and MassLynx respectively. Samples were analysed in a range of solvents and data measured in the mass range m/z 250.1 to 1478.6 and the CCS range 194.9 Ã 2 to 368.5 Ã 2. Although it was not possible to obtain data for high size and m/z dendrimers, the reproducibility of the data obtained suggests that SpheriCalÂ® compounds could form the basis of ion mobility calibrants in the future. Proteins were then analysed to determine the reproducibility of CCS data when compared to rigid dendrimers. Two small monomeric proteins, ubiquitin and Î±-synuclein were investigated. Ubiquitin is a regulatory protein found in eukaryotic organisms that has a mass of 8.6 kDa, before analysis it was denatured to promote a range of extended conformers, by making a wider range of charge sites accessible, ubiquitin7+ to ubiquitin13+ were analysed. Î±-synuclein is a flexible protein of mass 14.5 kDa, which was investigated to determine factors that may affect the conformational range observed as it forms aggregates. Mass spectra and arrival time distributions show substantial conformational change to Î±-synuclein over 24h at 37 oC in ammonium acetate. The effect of ionic strength of solution on the observed conformations of Î±-synuclein was examined.
|Date of Award||1 Aug 2020|
|Supervisor||Konstantin Novoselov (Supervisor) & Perdita Barran (Supervisor)|
- Collisional Cross Section
- Ion Mobility