Charge Site Mass Spectra: Conformation-Sensitive Components of the Electron Capture Dissociation Spectrum of a Protein
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A conventional electron capture dissociation (ECD) spectrum of a protein is uniquely characteristic of the first dimension of its linear structure. This sequence information is indicated by summing the primary c m+ and z m+• products of cleavage at each of its molecular ion’s inter-residue bonds. For example, the ECD spectra of ubiquitin (M + nH)n+ ions, n = 7–13, provide sequence characterization of 72 of its 75 cleavage sites from 1843 ions in seven c (1–7)+ and eight z (1–8)+• spectra and their respective complements. Now we find that each of these c/z spectra is itself composed of “charge site (CS)” spectra, the c m+ or z m+• products of electron capture at a specific protonated basic residue. This charge site has been H-bonded to multiple other residues, producing multiple precursor ion forms; ECD at these residues yields the multiple products of that CS spectrum. Closely similar CS spectra are often formed from a range of charge states of ubiquitin and KIX ions; this indicates a common secondary conformation, but not the conventional α-helicity postulated previously. CS spectra should provide new capabilities for comparing regional conformations of gaseous protein ions and delineating ECD fragmentation pathways.
Key wordsElectron capture dissociation (ECD) Charge site (CS) mass spectra Protein ion conformation
The authors acknowledge funding by the Austrian Science Fund (FWF, Y372 to K.B.) and the General Medical Institute of the National Institutes of Health (GM16609 to F.W.M.).
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