Mechanospray Ionization MS of Proteins Including in the Folded State and Polymers

• Published in: Journal of the American Society for Mass Spectrometry Vol. 33; no. 5; pp. 772 - 782
• Main Authors: Dugan, Liam D, Bier, Mark E
• Format: Journal Article
• Language: English
• Published: United States American Chemical Society 04-05-2022
• Subjects: Heme; Ions - chemistry; Polymers; Proteins - chemistry; Spectrometry, Mass, Electrospray Ionization - methods; folded proteins; PEG; proteins; PPG; low internal energy; native MS; peptides; mechanospray ionization
• ISSN: 1044-0305; 1879-1123
• DOI: 10.1021/jasms.1c00344

Mechanospray ionization (MoSI) is a technique that produces ions directly from solution-like electrospray ionization (ESI) but without the need of a high voltage. In MoSI, mechanical vibrations aerosolize solution phase analytes, whereby the resulting microdroplets can be directed into the inlet orifice of a mass spectrometer. In this work, MoSI is applied to biomolecules up to 80 kDa in mass in both denatured and native conditions as well as polymers up to 12 kDa in mass. The various MoSI devices used in these analyses were all comprised of a piezoelectric annulus attached to a central metallic disk containing an array of 4 to 7 μm diameter holes. The devices vibrated in the 100–170 kHz range to generate a beam of microdroplets that ultimately resulted in ion formation. A linear quadrupole ion trap (LIT) and orbitrap mass spectrometer were used in the analysis to investigate higher mass proteins at both native (folded) and denatured (unfolded) conditions. MoSI native mass spectra of proteins acquired on the orbitrap and LIT instrument demonstrated that proteins could remain intact and in a folded state. In the case of native MS of holomyoglobin, the intact folded protein remained mostly bound noncovalently to the heme group, and typically, the spectra showed reduced loss of the heme group by MoSI as compared to ESI. In both non-native and native protein analyses examples, broader often multimodal distributions to lower charge states were observed. When using the LIT instrument, a significant increase in the relative abundance of dimers was observed by MoSI as compared to ESI. The softness of the MoSI technique was evidenced by the lack of fragmentation, the formation of dimers as also noted by others ( J. Mass Spectrom. 2016, 424–429 ) and under native conditions, the retention of proteins in one or more presumed folded structures and for holomyoglobin the high retention of the heme group. When analyzing polyethylene glycol (PEG) and polypropylene glycol (PPG), MoSI also generated a broader distribution to lower charge states than ESI. By using the improved separation of peaks at lower charge states and all the charge states available, MoSI data should provide an improved ionization method to obtain more accurate mass and dispersity values for some polymers.