Electrospray ion mobility spectrometry of intact viruses

Thomas, John J.; Bothner, Brian; Traina, Joe; Benner, W. Henry; Siuzdak, Gary

doi:10.1155/2004/376572

Cited by 82 publications

(74 citation statements)

References 25 publications

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“…ES-DMA can achieve this objective by correlating identity with size. Thomas et al, [35] has shown that the components of mixtures of very similar sized viruses can be distinguished based on size by ES-DMA.…”

Section: Discussion and Summarymentioning

confidence: 99%

Electrospray–differential mobility analysis of bionanoparticles

Guha

Tarlov

et al. 2012

Trends in Biotechnology

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The growth of the multibillion dollar bionanoparticle industry has spurred the development of new physical characterization methods. One such method, electrospraydifferential mobility analysis (ES-DMA) constitutes an electrospray for aerosolization of bionanoparticles (such as viruses, gold-nanoparticles, proteins, nanoparticle-protein complexes) and an ion mobility method that operates at atmospheric conditions, and separates bionanoparticles spatially. This dissertation identifies some relevant "problem" areas for ES-DMA by reviewing selected applications.Some such problems are: proteins while passing through ES capillaries are found to interact with it and thus produce time dependent size distributions. Further, it is thought that adsorbed proteins may subsequently desorb and influence size distributions with the ES-DMA which may concomitantly affect quantification of aggregates. These artifacts are studied systematically and it is demonstrated that ES-DMA can quantify adsorption-desorption of complex protein mixtures at high shear rates. Further, it is shown that desorbing proteins do not have a significant effect on size distributions. Another artifact of the ES takes place during the aersolization process. Two units (called monomers) of a bionanoparticle may get encapsulated in the same ES droplet and upon drying of the droplet create artificial dimers thus affecting quantification with ES-DMA. Assuming Poisson distribution, this thesis provides a systematic approach that can be undertaken to eliminate this artifact. A third artifact arises from the low sensitivity of the DMA to size increase. When a ligand (for e.g. protein) adsorbs to a bionanoparticle it creates an increase in the size of the later, which can be used to quantify the amount of ligand adsorbed per bionanoparticle. As ligands can change conformations upon adsorption, using ES-DMA for such applications may be flawed. This issue has been identified and a solution has been provided by integrating a mass analyzer after the ES-DMA.After correcting for these artifacts, this dissertation delves into characterization of different types of bionanoparticles and demonstrates that ES-DMA has several advantages over other traditional techniques such as transmission electron microscopy, size exclusion chromatography, analytical ultracentrifugation, dynamic light scattering and plaque assay and thus has immense potential to become a process analytical technique in biomanufacturing environments. ELECTROSPRAY-DIFFERENTIAL MOBILITY ANALYSIS OF BIONANOPARTICLES

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Section: Discussion and Summarymentioning

confidence: 99%

Electrospray–differential mobility analysis of bionanoparticles

Guha

Tarlov

et al. 2012

Trends in Biotechnology

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“…ESI-GEMMA studies of primarily globular proteins have shown a correlation between electrophoretic diameter and relative molecular mass for proteins and protein complexes ranging from 3 kDa to 2 MDa and measuring 2.6 to 21.7 nm in EMD [8]. Highlighting the role of water in supramolecular complexes and the potential of ESI-GEMMA as a structural elucidation tool, Thomas et al showed that the measured electrophoretic mobility for selected viruses were within 15% of the diameters measured by X-ray crystallographic studies [13]. Furthermore, studies using other ESI-IMS techniques have shown agreement between X-ray crystal structure and gas-phase structure dimensions for both globular proteins and large protein complexes.…”

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confidence: 97%

“…tions [3,7,8,13]. Within the electrospray unit of the GEMMA apparatus, the protein solution is sprayed into a mist of highly charged droplets, which are then charge-reduced by a bipolar neutralizer.…”

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confidence: 99%

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Sizing large proteins and protein complexes by electrospray ionization mass spectrometry and ion mobility

Kaddis

Lomeli

Yin

et al. 2007

J. Am. Soc. Mass Spectrom.

147

149

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Mass spectrometry (MS) and ion mobility with electrospray ionization (ESI) have the capability to measure and detect large noncovalent protein-ligand and protein-protein complexes. Using an ion mobility method of gas-phase electrophoretic mobility molecular analysis (GEMMA), protein particles representing a range of sizes can be separated by their electrophoretic mobility in air. Highly charged particles produced from a protein complex solution using electrospray can be manipulated to produce singly charged ions, which can be separated and quantified by their electrophoretic mobility. Results from ESI-GEMMA analysis from our laboratory and others were compared with other experimental and theoretically determined parameters, such as molecular mass and cryoelectron microscopy and X-ray crystal structure dimensions. There is a strong correlation between the electrophoretic mobility diameter determined from GEMMA analysis and the molecular mass for protein complexes up to 12 MDa, including the 93 kDa enolase dimer, the 480 kDa ferritin 24-mer complex, the 4.6 MDa cowpea chlorotic mottle virus (CCMV), and the 9 MDa MVP-vault assembly. ESI-GEMMA is used to differentiate a number of similarly sized vault complexes that are composed of different N-terminal protein tags on the MVP subunit. The average effective density of the proteins and protein complexes studied was 0.6 g/cm 3 . Moreover, there is evidence that proteins and protein complexes collapse or become more compact in the gas phase in the absence of water. (J Am Soc Mass Spectrom 2007, 18, 1206 -1216

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“…Recent advances have seen its successful application to systems, including the ribosome [1], intact viruses [2,3], membrane associated protein assemblies [4,5], and various protein-small molecule ligand complexes [6,7]. The use of ESI-MS to study noncovalent protein interactions offers several advantages over other biophysical techniques.…”

Section: Introductionmentioning

confidence: 99%

Evidence for the Preservation of Native Inter- and Intra-Molecular Hydrogen Bonds in the Desolvated FK-Binding Protein·FK506 Complex Produced by Electrospray Ionization

Hopper

Rawlings

Afonso

et al. 2012

J. Am. Soc. Mass Spectrom.

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It is now well established that electrospray ionization (ESI) is capable of introducing noncovalent protein assemblies into a desolvated environment, thereby allowing their analysis by mass spectrometry. The degree to which native interactions from the solution phase are preserved in this environment is less clear. Site-directed mutagenesis of FK506-binding protein (FKBP) has been employed to probe specific intra-and inter-molecular interactions within the complex between FKBP and its ligand FK506. Collisional activation of wild-type and mutant-FKBP•FK506 ions, generated by ESI, demonstrated that removal of native protein-ligand interactions formed between residues Asp37, Tyr82, and FK506 significantly destabilized the complex. Mutation of Arg42 to Ala42, or Tyr26 to Phe26 also resulted in lower energy dissociation of the FKBP·FK506 complex. Although these residues do not form direct H-bonds to FK506, they interact with Asp37, ensuring its correct orientation to associate with the ligand. Comparison with solution-based affinity measurements of these mutants has been discussed, including the stabilization afforded by ordered water molecules. Ion mobility spectrometry (IMS) has been employed to provide gas-phase structural information on the unfolding of the complexes. The [M + 6H] 6+ complexes of the wild-type and mutants have been shown to resist unfolding and retain compact conformations. However, removal of the basic Arg42 residue was found to induce significant structural weakening of the [M + 7H] 7+ complex when raised to dissociation-level energies. Overall, destabilization of the FKBP·FK506 complex, resulting from targeted removal of specific H-bonds, provides evidence for the preservation of these interactions in the desolvated wild-type complex.

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Electrospray ion mobility spectrometry of intact viruses

Cited by 82 publications

References 25 publications

Electrospray–differential mobility analysis of bionanoparticles

Electrospray–differential mobility analysis of bionanoparticles

Sizing large proteins and protein complexes by electrospray ionization mass spectrometry and ion mobility

Evidence for the Preservation of Native Inter- and Intra-Molecular Hydrogen Bonds in the Desolvated FK-Binding Protein·FK506 Complex Produced by Electrospray Ionization

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