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J Am Soc Mass Spectrom. 2010 Jan;21(1):127-31. doi: 10.1016/j.jasms.2009.09.014. Epub 2009 Sep 30.

New reagents for increasing ESI multiple charging of proteins and protein complexes.

Author information

1
Department of Chemistry and Biochemistry, David Geffen School of Medicine, University of California-Los Angeles, Los Angeles, California, USA.

Abstract

The addition of m-nitrobenzyl alcohol (m-NBA) was shown previously (Lomeli et al., J. Am. Soc. Mass Spectrom. 2009, 20, 593-596) to enhance multiple charging of native proteins and noncovalent protein complexes in electrospray ionization (ESI) mass spectra. Additional new reagents have been found to "supercharge" proteins from nondenaturing solutions; several of these reagents are shown to be more effective than m-NBA for increasing positive charging. Using the myoglobin protein-protoporphyrin IX (heme) complex, the following reagents were shown to increase ESI charging: benzyl alcohol, m-nitroacetophenone, m-nitrobenzonitrile, o-NBA, m-NBA, p-NBA, m-nitrophenyl ethanol, sulfolane (tetramethylene sulfone), and m-(trifluoromethyl)-benzyl alcohol. Based on average charge state, sulfolane displayed a greater charge increase (61%) than m-NBA (21%) for myoglobin in aqueous solutions. The reagents that promote higher ESI charging appear to have low solution-phase basicities and relatively low gas-phase basicities, and are less volatile than water. Another feature of mass spectra from some of the active reagents is that adducts are present on higher charge states, suggesting that a mechanism by which proteins acquire additional charge involves direct interaction with the reagent, in addition to other factors such as surface tension and protein denaturation.

PMID:
19854660
PMCID:
PMC2821426
DOI:
10.1016/j.jasms.2009.09.014
[Indexed for MEDLINE]
Free PMC Article

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