Structural characterization of glycopeptides by N-terminal protein ladder sequencing.
High-sensitivity and high-throughput mass spectrometry (MS) has become an important tool for characterizing glycopeptides. Here, we analyzed synthetic O-linked glycopeptides using matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) MS. First, we applied MALDI-quadrupole ion trap (QIT)-TOF MS, which enables collision-induced dissociation-MSn analysis for fine structural characterization. Subsequent MS/MS of sodium adduct ions selected as precursor ions yielded detailed information about the site of oligosaccharide attachment as well as the carbohydrate and amino acid sequences; however, these MS/MS spectra were very complex. To obtain easily interpretable and simple spectra, we used N-terminal protein ladder sequencing coupled with MALDI-TOF MS. From the extremely simple resulting spectra, we were able to determine the glycosylation sites, amino acid sequences, and oligosaccharide molecular weights of the glycopeptides.
Sphingolipid Expression Laboratory, Supra Biomolecular System Research Group, RIKEN Frontier Research System, Wako-shi, Saitama 351-0198, Japan.
Yusuke
Y
Suzuki
Minoru
M
Suzuki
Yoshiaki
Y
Nakahara
Yukishige
Y
Ito
Emi
E
Ito
Naoko
N
Goto
Kozo
K
Miseki
Junko
J
Iida
Akemi
A
Suzuki
D000595:Amino Acid Sequence; D001665:Binding Sites; D006020:Glycopeptides / Q000032:analysis* / Q000737:chemistry*; D006031:Glycosylation; D006868:Hydrolysis; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009844:Oligosaccharides / Q000737:chemistry; D015203:Reproducibility of Results; D020539:Sequence Analysis, Protein / Q000379:methods*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods*
D006020:Glycopeptides; D009844:Oligosaccharides