Increasing Protein Charge State When Using Laser Electrospray Mass Spectrometry

Abstract

Abstract. Femtosecond (fs) laser vaporization is used to transfer cytochrome c, myoglobin, lysozyme, and ubiquitin from the condensed phase into an electrospray (ES) plume consisting of a mixture of a supercharging reagent, m-nitrobenzyl alcohol (m-NBA), and trifluoroacetic acid (TFA), acetic acid (AA), or formic acid (FA). Interaction of acid-sensitive proteins like cytochrome c and myoglobin with the highly charged ES droplets resulted in a shift to higher charge states in comparison with acid-stable proteins like lysozyme and ubiquitin. Laser electrospray mass spectrometry (LEMS) measurements showed an increase in both the average charge states (Z avg ) and the charge state with maximum intensity (Z mode ) for acid-sensitive proteins compared with conventional electrospray ionization mass spectrometry (ESI-MS) under equivalent solvent conditions. A marked increase in ion abundance of higher charge states was observed for LEMS in comparison with conventional electrospray for cytochrome c (ranging from 19+ to 21+ versus 13+ to 16+) and myoglobin (ranging from 19+ to 26+ versus 18+ to 21+) using an ES solution containing m-NBA and TFA. LEMS measurements as a function of electrospray flow rate yielded increasing charge states with decreasing flow rates for cytochrome c and myoglobin.

Classification of Gunshot Residue Using Laser Electrospray Mass Spectrometry and Offline Multivariate Statistical Analysis

“…LEMS has been used to detect and classify different types of explosives, such as inorganic salts, high explosives, and unburnt smokeless powders . Furthermore, LEMS analyses have been conducted on a variety of analytes including pharmaceuticals, lipids, narcotics, benzylpyridinium molecules, dipeptides, proteins, − plant, , and mammalian tissue suggesting LEMS would be amenable for GSR analysis.…”

Enhanced-Fluidity Liquid Chromatography–Mass Spectrometry for Intact Protein Separation and Characterization

“…According to Fenn’s model of ion formation, which is an extension of the ion evaporation model, the charges move closer together to give an increased surface charge density as the droplet evaporates . When the smaller droplets are formed in the early stage of the ESI process, a protein will spend a longer time in the vicinity of the high surface charge density leading to higher charge states . In addition to the increased surface charge density, the lower surface tension requires fewer Coulombic fission events to create gas phase ions, resulting in higher ionization efficiency …”

Laser Desorption/Ablation Postionization Mass Spectrometry: Recent Progress in Bioanalytical Applications

“…Applications of LEMS in biological analysis have been focused on characterization of small metabolites and macro biomolecules including enzyme, proteins, and phosphorous lipids from human blood and egg white/yolk (Judge, Brady, & Levis, 2010); amphiphilic lipids and hydrophobic proteins deposited on steel and glass substrates (Brady, Judge, & Levis, 2011a); plant tissue typing using compressive linear classification (Judge et al, 2011); plant phenotype discrimination using multivariate statistical analysis (Flanigan et al, 2012); acid sensitive proteins with increased charge states when infusing supercharging reagents into the electrospray or deceasing the electrospray flow rate (Karki et al, 2015); folded proteins and hemoglobin α subunit‐heme complex from whole blood with LEMS integrating with a fiber‐based femtosecond laser at 1042 nm (Shi et al, 2015); and small metabolites from plant samples using the low pulse energy fiber‐based laser for analytes desorption, with comparable results to high energy Ti: Sapphire laser‐based LEMS measurements (Shi et al, 2016b).…”