TY - JOUR
T1 - New reagents for enhanced liquid chromatographic separation and charging of intact protein ions for electrospray ionization mass spectrometry
AU - Valeja, Santosh G.
AU - Tipton, Jeremiah D.
AU - Emmett, Mark R.
AU - Marshall, Alan G.
PY - 2010/9/1
Y1 - 2010/9/1
N2 - Electrospray ionization produces multiply charged ions, thereby lowering the mass-to-charge ratio for peptides and small proteins to a range readily accessed by quadrupole ion trap, orbitrap, and ion cyclotron resonance (ICR) mass analyzers (m/z = 400-2000). For Fourier transform mass analyzers (orbitrap and ICR), higher charge also improves signal-to-noise ratio, mass resolution, and mass accuracy. Addition of m-nitrobenzyl alcohol (m-NBA) or sulfolane has previously been shown to increase the charge states of proteins. Moreover, polar aprotic dimethylformamide (DMF) improves chromatographic separation of proteolytic peptides for mass analysis of solution-phase protein hydrogen/deuterium exchange for improved (78-96%) sequence coverage. Here, we show that addition of each of the various modifiers (DMF, thiodiglycol, dimethylacetamide, dimethylsulfoxide, and N-methylpyrrolidone) can significantly increase the charge states of proteins up to 78 kDa. Moreover, incorporation of the same modifiers into reversed-phase liquid chromatography solvents improves sensitivity, charging, and chromatographic resolution for intact proteins.
AB - Electrospray ionization produces multiply charged ions, thereby lowering the mass-to-charge ratio for peptides and small proteins to a range readily accessed by quadrupole ion trap, orbitrap, and ion cyclotron resonance (ICR) mass analyzers (m/z = 400-2000). For Fourier transform mass analyzers (orbitrap and ICR), higher charge also improves signal-to-noise ratio, mass resolution, and mass accuracy. Addition of m-nitrobenzyl alcohol (m-NBA) or sulfolane has previously been shown to increase the charge states of proteins. Moreover, polar aprotic dimethylformamide (DMF) improves chromatographic separation of proteolytic peptides for mass analysis of solution-phase protein hydrogen/deuterium exchange for improved (78-96%) sequence coverage. Here, we show that addition of each of the various modifiers (DMF, thiodiglycol, dimethylacetamide, dimethylsulfoxide, and N-methylpyrrolidone) can significantly increase the charge states of proteins up to 78 kDa. Moreover, incorporation of the same modifiers into reversed-phase liquid chromatography solvents improves sensitivity, charging, and chromatographic resolution for intact proteins.
UR - http://www.scopus.com/inward/record.url?scp=77956256592&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=77956256592&partnerID=8YFLogxK
U2 - 10.1021/ac1016858
DO - 10.1021/ac1016858
M3 - Article
C2 - 20704305
AN - SCOPUS:77956256592
SN - 0003-2700
VL - 82
SP - 7515
EP - 7519
JO - Analytical Chemistry
JF - Analytical Chemistry
IS - 17
ER -