Studying The Effect Of Alkali Metal On The Dissociation Of Polypeptides In Gas Phase By Using Electrospray Ionization Mass Spectrometry

In recent years, mass spectrometry technology, especially electrospray ionization (ESI) and matrix-assisted laser desorption ionization (MALDI) and other "soft" ionization mass spectrometry technologies, has not only used to detect the heat-labile, strong polar, hard-volatilization compounds, but also allow complex completely enter the gas phase from the solution phase to be detected directly under certain conditions. Thus we could obtain information of the stoichiometry of molecular interactions, combined with the binding situation and the information of energy under near physiological state, so that the application of mass spectrometry has been extended from small molecule to large biological molecules, widely being used in peptide sequencing and protein structure analysis. By using Electrospray ionization mass spectrometry(ESI-MS) to study small molecules-peptides gas chemistry, one can exclude the interference of the solvation effect, revealing the inherent binding nature of small molecule -peptide, and thus the analysis of peptides and proteins has many advantages. In this dissertation, we make use of ESI-MS as the main research technique to investigate fragmentation reactions of non-covalent complex of glutathione and amino acids and the effect of alkali metal on the dissociation of polypeptides in gas phase.First we explore the fragmentation reactions of non-covalent complex of glutathione (GSH) and amino acids, to investigate the main factors affecting the fragmentation process. The secondary tandem mass spectrometry (MS2) results indicated that collision gas thickness (CGT) and collision energy are two important conditions affecting the fragmentation pathway of glutathione complex. When the collision gas thickness maintained at 50x 1013 molecules/cm2, increasing collision energy from 5 eV to 80 eV, the dissociation of His-GSH complex took place, leading to the formation of [His+H]+ and [GSH+H]+. However, it can be seen that no further dissociation of GSH occurred and the fragmentation of noncovalent bond was the main pathway of the complex. When CGT rise up to 75×1013 molecules/cm2, the covalent bond also began to dissociate, leading to the formation of y2 and b2 for glutathione. Such phenomena could be also observed in Lys-GSH complex. To confirm the fragment ion y2 of glutathione, MS3 was also performed and y1 was detected.Then we use electrospray ionization linear ion trap mass spectrometry to study the gas phase chemical interactions of alkali metal lithium and three pentapeptides GGGGG, KKKKK and SSSSS, with collision induced dissociation of the protonated pentapeptide, single lithiation and double lithiation pentapeptide. Experimental results show that the dossociation reation results of single lithiation pentapeptide and protonated pentapeptides was near in the type and relative abundance of product ions, etc, suggesting that they experience similar dissociation pathway, so it has proved that the impact of lithium on the dissciation of polypeptides is not significant. When two peptides combined with lithium, the dissociation results were much different from single lithiation and double lithition pentapeptides, indicating that two lithium had greatly influenced the performance of peptides dissociation, the reason can attribute to the interaction of two lithium and peptides can activate the polypeptide, inducing its cleavage in more sites, resulting in more abundant sequence ions information, so as to be a complementary method for protonated peptides sequencing.And at last, we synthetically studied the gas phase chemical interactions of alkali metal sodium, potassium, rubidium and three pentapeptides GGGGG, KKKKK and SSSSS, with low energy collision induced dissociation of the protonated pentapeptide, single alkali metalation and double alkali metalation pentapeptide. By studying the fragment ions distribution, the dissociation differences among three pentapeptide complexes were qualitatively compared, and also explored the effects of different alkali metals on the peptides dissociation. Experiment results showed that, to all the alkali metals, when a single alkali metal binds to a pentapeptide, the dissociation outcome differs from the protonated peptide in product ions distribution, indicating that all the single alkali metals can affect the dissociation patterns of peptides from a certain extent. The smaller the radius of the alkali metal is, the less impact it will have, and the product ions distribution are more alike that of the protonated peptide. While the impact of two alkali metals on the dissociation patterns depends on the character of the peptides. When there is no side chain groups on the peptides, the combination of two alkali metals and the peptide will further improve the activation of peptide, inducing it to dissociate in more sites, whether this activatation cleavage is direct or indirect, it will lead to the types and numbers of product ions both increase. Due to the similarity among the relative abundance of product ions, the difference between the radius of alkali metals and affinity among peptides are regarded as disappeared. But to those peptides with side chains, the interaction of the two alkali metals and peptides would affect the dissociation patterns of peptides, and the degree of influence depends on the affinity between alkali metals and peptides, radius of alkali metals, characteristics of side chains and many other factors.