Quantitative analysis of modified proteins and their positional isomers by tandem mass spectrometry: human histone H4.
Here we show that fragment ion abundances from dissociation of ions created from mixtures of multiply modified histone H4 (11 kDa) or of N-terminal synthetic peptides (2 kDa) correspond to their respective intact ion abundances measured by Fourier transform mass spectrometry. Isomeric mixtures of modified forms of the same protein are resolved and quantitated with a precision of </=5% using the relative ratios of their fragment ions, with intact protein ions created by electrospray greatly easing many of the systematic biases that more strongly affect small peptides (e.g., differences in ionization efficiency and ion m/z values). The ion fragmentation methods validated here are directly extensible to intact human proteins to derive quantitative information on the highly related and often isomeric protein forms created by combinatorial arrays of posttranslational modifications.
Pesavento
James J
JJ
Center for Biophysics and Computational Biology, Department of Cell and Developmental Biology, Institute for Genomic Biology (IGB), University of Illinois at Urbana-Champaign, Urbana, Illinois 61801, USA.
Mizzen
Craig A
CA
Kelleher
Neil L
NL
D002851:Chromatography, High Pressure Liquid; D006367:HeLa Cells; D006657:Histones / Q000032:analysis* / Q000737:chemistry; D006801:Humans; D020033:Protein Isoforms / Q000032:analysis* / Q000737:chemistry; D011994:Recombinant Proteins / Q000032:analysis / Q000737:chemistry; D053719:Tandem Mass Spectrometry / Q000379:methods*
D006657:Histones; D020033:Protein Isoforms; D011994:Recombinant Proteins