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Exploring the Mechanism of Salt-Induced Signal Suppression in Protein Electrospray Mass Spectrometry Using Experiments and Molecular Dynamics Simulations
journal contribution
posted on 2015-02-17, 00:00 authored by Haidy Metwally, Robert G. McAllister, Lars KonermannProtein analyses by electrospray
ionization (ESI) mass spectrometry
can suffer from interferences caused by nonvolatile salts. The mechanistic
basis of this effect remains to be fully investigated. In the current
work we explore the behavior of proteins under native and denaturing
conditions in the presence of NaCl, CsCl, and tetrabutyl ammonium
chloride (NBu4Cl). All three salts interfere with the formation
of “clean” [M + zH]z+ protein ions by progressively deteriorating spectral S/N
ratios. We propose that salt interferences can be dissected into two
independent aspects, i.e., (i) peak splitting by adduct formation
and (ii) protein ion suppression. NaCl degrades the spectral quality
by forming heterogeneous [M + zH + n(Na – H) + m(Cl + H)]z+ ions, while the integrated protein ion intensity remains
surprisingly robust. Conversely, NBu4Cl does not cause
any adduction, while dramatically reducing the protein ion yield.
These findings demonstrate that adduct formation and protein ion suppression
are indeed unrelated effects that may occur independently of one another.
Other salts, such as CsCl, can give rise to a combination of the two
scenarios. Molecular dynamics simulations of water droplets charged
with either Na+ or NBu4+ provide
insights into the mechanism underlying the observed effects. Na+ containing droplets evolve relatively close to the Rayleigh
limit (z/zR ≈
0.74), whereas the z/zR values of NBu4+ charged droplets are considerably
lower (∼0.59). This difference is due to the high surface affinity
of NBu4+, which facilitates charge ejection
from the droplet. We propose that the low z/zR values encountered in the presence of NBu4+ suppress the Rayleigh fission of parent droplets
in the ESI plume, thereby reducing the yield of progeny droplets that
represent the precursors of gaseous protein ions. In addition, the
rate of solvent evaporation is reduced in the presence of NBu4+. Both of these factors lower the protein signal
intensity. NaCl does not interfere with droplet fission, such that
protein ions continue to form with high yieldalbeit in heavily
adducted form. Our findings expand on earlier proposals of charge
competition as a key factor during the ESI process for salt-contaminated
solutions.