| Autor: |
Weiss VU; Institute of Chemical Technologies and Analytics, TU Wien , Getreidemarkt 9/164, A-1060 Vienna, Austria., Bereszcazk JZ; Bijvoet Centre for Biomolecular Research and Utrecht Institute of Pharmaceutical Sciences, Utrecht University , NL-3584 CH Utrecht, The Netherlands., Havlik M; Institute of Chemical Technologies and Analytics, TU Wien , Getreidemarkt 9/164, A-1060 Vienna, Austria., Kallinger P; Faculty of Physics, University of Vienna , A-1090 Vienna, Austria., Gösler I; Department of Medical Biochemistry, Max F. Perutz Laboratories, Medical University of Vienna, Vienna Biocenter (VBC) , A-1030 Vienna, Austria., Kumar M; Department of Medical Biochemistry, Max F. Perutz Laboratories, Medical University of Vienna, Vienna Biocenter (VBC) , A-1030 Vienna, Austria., Blaas D; Department of Medical Biochemistry, Max F. Perutz Laboratories, Medical University of Vienna, Vienna Biocenter (VBC) , A-1030 Vienna, Austria., Marchetti-Deschmann M; Institute of Chemical Technologies and Analytics, TU Wien , Getreidemarkt 9/164, A-1060 Vienna, Austria., Heck AJ; Bijvoet Centre for Biomolecular Research and Utrecht Institute of Pharmaceutical Sciences, Utrecht University , NL-3584 CH Utrecht, The Netherlands., Szymanski WW; Faculty of Physics, University of Vienna , A-1090 Vienna, Austria., Allmaier G; Institute of Chemical Technologies and Analytics, TU Wien , Getreidemarkt 9/164, A-1060 Vienna, Austria. |
| Abstrakt: |
Gas-phase electrophoretic mobility molecular analysis (GEMMA) separates nanometer-sized, single-charged particles according to their electrophoretic mobility (EM) diameter after transition to the gas-phase via a nano electrospray process. Electrospraying as a soft desorption/ionization technique preserves noncovalent biospecific interactions. GEMMA is therefore well suited for the analysis of intact viruses and subviral particles targeting questions related to particle size, bioaffinity, and purity of preparations. By correlating the EM diameter to the molecular mass (Mr) of standards, the Mr of analytes can be determined. Here, we demonstrate (i) the use of GEMMA in purity assessment of a preparation of a common cold virus (human rhinovirus serotype 2, HRV-A2) and (ii) the analysis of subviral HRV-A2 particles derived from such a preparation. (iii) Likewise, native mass spectrometry was employed to obtain spectra of intact HRV-A2 virions and empty viral capsids (B-particles). Charge state resolution for the latter allowed its Mr determination. (iv) Cumulatively, the data measured and published earlier were used to establish a correlation between the Mr and EM diameter for a range of globular proteins and the intact virions. Although a good correlation resulted from this analysis, we noticed a discrepancy especially for the empty and subviral particles. This demonstrates the influence of genome encapsulation (preventing analytes from shrinking upon transition into the gas-phase) on the measured analyte EM diameter. To conclude, GEMMA is useful for the determination of the Mr of intact viruses but needs to be employed with caution when subviral particles or even empty viral capsids are targeted. The latter could be analyzed by native MS. |
