Supplemental activation method for high-efficiency electron-transfer dissociation of doubly protonated peptide precursors.
Electron-transfer dissociation (ETD) delivers the unique attributes of electron capture dissociation to mass spectrometers that utilize radio frequency trapping-type devices (e.g., quadrupole ion traps). The method has generated significant interest because of its compatibility with chromatography and its ability to: (1) preserve traditionally labile post-translational modifications (PTMs) and (2) randomly cleave the backbone bonds of highly charged peptide and protein precursor ions. ETD, however, has shown limited applicability to doubly protonated peptide precursors, [M + 2H]2+, the charge and type of peptide most frequently encountered in "bottom-up" proteomics. Here we describe a supplemental collisional activation (CAD) method that targets the nondissociated (intact) electron-transfer (ET) product species ([M + 2H]+*) to improve ETD efficiency for doubly protonated peptides (ETcaD). A systematic study of supplementary activation conditions revealed that low-energy CAD of the ET product population leads to the near-exclusive generation of c- and z-type fragment ions with relatively high efficiency (77 +/- 8%). Compared to those formed directly via ETD, the fragment ions were found to comprise increased relative amounts of the odd-electron c-type ions (c+*) and the even-electron z-type ions (z+). A large-scale analysis of 755 doubly charged tryptic peptides was conducted to compare the method (ETcaD) to ion trap CAD and ETD. ETcaD produced a median sequence coverage of 89%-a significant improvement over ETD (63%) and ion trap CAD (77%).
Swaney
Danielle L
DL
Departments of Chemistry and Biomolecular Chemistry, University of Wisconsin, Madison, Wisconsin 53706, USA.
McAlister
Graeme C
GC
Wirtala
Matthew
M
Schwartz
Jae C
JC
Syka
John E P
JE
Coon
Joshua J
JJ
D000595:Amino Acid Sequence; D004213:Dissociative Disorders; D004579:Electron Transport; D013058:Mass Spectrometry / Q000379:methods*; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000032:analysis*; D011506:Proteins / Q000737:chemistry*; D040901:Proteomics / Q000379:methods*; D011522:Protons; D014357:Trypsin / Q000737:chemistry
D010446:Peptide Fragments; D011506:Proteins; D011522:Protons; D014357:Trypsin