Analysis of a Common Cold Virus and Its Subviral Particles by Gas-Phase Electrophoretic Mobility Molecular Analysis and Native Mass Spectrometry

Abstract

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.

Size and molecular weight determination of polysaccharides by means of nano electrospray gas‐phase electrophoretic mobility molecular analysis (nES GEMMA)

“…GEMMA derived EMD/MW correlations for proteins, DNA and PEG have already been described . Exemplary data for proteins (own data), DNA (data from Mouradian et al .…”

“…Wasiak and colleagues employed gas‐phase electrophoresis for the analysis of dextran‐based nanoparticles as had Szymanski and co‐workers much earlier for a small dextran of 11.5 kDa. In the current work, we focus on the application of GEMMA for polysaccharide measurements in a more general approach; we intended to setup an EMD/MW correlation for the wide range of chemically divergent polysaccharides as previously done for proteins and DNA .…”

Acquiring Structural Information on Virus Particles with Charge Detection Mass Spectrometry

“…The particles are so massive and so heterogeneous (a continuous distribution of masses from 5 to at least 15 MDa) that conventional MS, as an ensemble measurement, could not conceivably extract masses from the m/z spectrum. Techniques such as gas-phase electrophoretic mobility molecular analysis (GEMMA) have been used to measure the gas-phase diameter of viral particles [64, 65, 68, 69] but a diameter spectrum of these samples would almost certainly be a broad, poorly resolved distribution, similar to Figure 3a and d. Similarly, an emerging single-particle mass spectrometry detector, a nanoelectromechanical oscillator, measures mass independently of charge [70].…”

Staying Alive: Measuring Intact Viable Microbes with Electrospray Ionization Mass Spectrometry

“…This technique has been used to detect intact MS2 bacteriophage at 24 ± 2 nm [30], and human rhinovirus with a diameter of 29.8 ± 0.3 nm [28]. Interestingly, since the measurements are based on the electrophoretic mobility (EM) measurements that are independent of known virus features, these results provide strong evidence that no large-scale disruption of tertiary or quaternary structure of the capsid occurred during ESI desolvation and ionization [31]. …”