High-energy electron transfer dissociation (HE-ETD) using alkali metal targets for sequence analysis of post-translational peptides.
Post-translational modifications (PTMs) of proteins are important in the activation, localization, and regulation of protein function in vivo. The usefulness of electron capture dissociation (ECD) and electron-transfer dissociation (ETD) in tandem mass spectrometry (MS/MS) using low-energy (LE) trap type mass spectrometer is associated with no loss of a labile PTM group regarding peptide and protein sequencing. The experimental results of high-energy (HE) collision induced dissociation (CID) using the Xe and Cs targets and LE-ETD were compared for doubly-phosphorylated peptides TGFLT(p)EY(p)VATR (1). Although HE-CID using the Xe target did not provide information on the amino acid sequence, HE-CID using the Cs target provided all the z-type ions without loss of the phosphate groups as a result of HE-ETD process, while LE-ETD using fluoranthene anion gave only z-type ions from z(5) to z(11). The difference in the results of HE-CID between the Xe and Cs targets demonstrated that HE-ETD process with the Cs target took place much more dominantly than collisional activation. The difference between HE-ETD using Cs targets and LE-ETD using the anion demonstrated that mass discrimination was much weaker in the high-energy process. HE-ETD was also applied to three other phosphopeptides YGGMHRQEX(p)VDC (2: X = S, 3: X = T, 4: X = Y). The HE-CID spectra of the doubly-protonated phosphopeptides (= [M + 2H](2+)) of 2, 3, and 4 using the Cs target showed a very similar feature that the c-type ions from c(7) to c(11) and the z-type ions from z(7) to z(11) were formed via N-C alpha bond cleavage without a loss of the phosphate group.
Hayakawa
Shigeo
S
Department of Chemistry, Graduate School of Science, Osaka Prefecture University, Sakai, Osaka, Japan. hayakawa@c.s.osakafu-u.ac.jp
Matsumoto
Shinya
S
Hashimoto
Mami
M
Iwamoto
Kenichi
K
Nagao
Hirofumi
H
Toyoda
Michisato
M
Shigeri
Yasushi
Y
Tajiri
Michiko
M
Wada
Yoshinao
Y
D002586:Cesium / Q000737:chemistry*; D004583:Electrons; D010748:Phosphopeptides / Q000032:analysis* / Q000378:metabolism; D010766:Phosphorylation; D011499:Protein Processing, Post-Translational; D040901:Proteomics / Q000379:methods*; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*; D014978:Xenon / Q000737:chemistry
D010748:Phosphopeptides; D002586:Cesium; D014978:Xenon