Characteristic Fragment Ions Of N-terminal Deuterohemin Hexapeptide Under Charge Remote Model In ESI-MS

With several years of development,electrospray mass spectrometry?ESI-MS?has been maturely applied to the detection and analysis of biological samples,because it is a kind of“soft ionization”,which can retain the complete structural information of the analyte molecules.In recent years,the birth of tandem mass spectrometry has made it a core technical method for proteomics.In tandem mass spectrometry,the fragmentation of peptides is regular,and trough the analysis of fragment ions,a lot of information about the structure of samples can be obtained.Therefore,it is particularly important to study the fragmentation rules of peptides in mass spectrometry.However,there is still a large blank in the research on the fragmentation mechanism waiting for us to fill.“Charge remote model”means that during the fragmentation reaction,the charge is fixed at a position far from the fragmentation site and does not directly participate in the fragmentation of the relevant chemical bond.Study on the formation of characteristic ions under"charge remote model"is of great significance for enriching the fragment ions information and improving the accuracy of protein sequencing.In this paper,N-terminal Deuterohemin Hexapeptide?DhHP-6?is used as research object.Several characteristic ions under charge remote model are found and explained through the series of experiments such as amino acid substitution,electrospray mass spectrometry and theoretical calculations.There are three chapters in this paper.And the main contents of each chapter are as follow:The first chapter is the introduction.In this chapter,we introduced the components of mass spectrometry,the theory of tandem mass spectrometry,and its application in proteomics in detail.Also,the currently known fragmentation mechanism of peptides under“free proton”model and“charge remote”model was summarized.Finally,we introduced the structural characteristics of DhHP-6 and its biological functions,and put forward the ideas and significance of this paper.The second chapter is the study of the characteristic b₂-CO₂ ions of DhHP-6 under collision-induced dissociation mode.Through amino acid substitution experiments,using mass spectrometry technology,combined with UV and other characterization methods,we proposed a new fragmentation mechanism,that in gas phase the unprotonated oxazolone b₂ ion containing the histidine side chain imidazole ring loses a molecule of CO₂ through the bond cleavage and hydrogen transfer,leading to the formation of a conjugated structure.The corresponding theoretical calculation results also proved that this process is feasible in energy.There are a lot number of proteins in nature that contain heme structures and form the proximal Fe³⁺-His coordination.The model peptides used in this work provide a new fragmentation pattern information.The third chapter is the work on the peptide derivatives of DhHP-6 containing chiral amino acids.The differentiation of chiral amino acids in this kind of peptides under charge remote model was studied.In the work of this chapter,we found that peptides containing N-terminal deuterohemin exhibited a special fragmentation method during the secondary fragmentation of the electrospray mass spectrometry experiment.Based on the special fragments,the Aspartic acid chiral D/L isomerism can be distinguished.D-Aspartic acid produces special[b₅-72]⁺ions,while L-Aspartic acid produces special[b₅-90]⁺ions.We analyzed the fragmentation of single-charged and double-charged precursor ions in tandem mass spectrometry,combining with theoretical calculations,and explained the formation of characteristic ions.We hope that the characteristic fragment ions mentioned in the paper could enrich the known charge remote model fragmentation mechanism and play a role in improving the accuracy of protein identification.