N-磷酰化多肽的质子加合峰与钠离子加合峰的多级质谱裂解规律的差异性研究.doc

  Fragmentation difference between the protonated and sodiated phosphonamidate peptide in multistage mass spectrometry# 5 10 15 20 25 30 35 40 Liu Yan, Chen Peiyan, Yuan Hang, Xu Pengxiang, Zhao Yufen* (Department of Chemical Biology, College of Chemistry and Chemical engineering, Xiamen University, FuJian XiaMen 361005) Abstract: In the present work, N-phosphorylation method with three phosphorus reagents had been used as a derivatization method for sequencing actin binding domain of thymosin by ESI-MS/MS. Through comparing the fragmentation mechanism of proton adducts with sodium adducts, it was observed that proton adducts could provide more fragmentation information than sodium adducts. Proton tends to be adducted on nitrogen atom of amide bonds, which induces amide bonds to be cleaved along amide bonds in peptides and lose easily an amino acid molecule from the C-terminal firstly. While, sodium ion is adducted on oxygen atom of carbonyl or hydroxyl group in peptides and made the phosphorylated peptides tend to lose amino acid residue from the C-terminal. Furthermore, phosphorylated actin binding domain of thymosin, the heptapeptide protected on N-terminal, was sequenced successfully by ESI-MS/MS, it is implied that phosphoryl derivatization sequencing method also suits for the peptides modified on N-terminal. Keywords: phosphorylated peptides; proton adducts; sodium adducts; electrospray ionization mass spectrometry; collision-induced dissociation 0 Introduction Primary sequence of protein is very important for studies of their structure-function relationship. Generally, it is determined by cleaving the protein into peptides with proteolytic enzyme or chemical reagent, and then using one or several methods to sequence the peptides. [1] Edman method [2] is widely used in peptide sequencing, but it does not suit for the peptides modified on N-terminal. Following the development of “soft” ionization technique such as FAB, MALDI and ESI, numerous mass spectrome