Theoretical Study On The Characteristic Dissociations Of The Phenylalanine Containing Peptide Radical Cations

The study of dissociation mechanisms of the protonated peptides is the basis of protein sequencing,and it is also one of the research focuses of mass spectrometry-based proteomics.It plays an important role in the exploration of protein structures and functions.Based on the density functional theory M06-2X/6-31++G(d,p)method,this dissertation selects phenylalanyltyrosine dipeptide radical cation[FY]^·+as the research object,in which MS/MS spectra a characteristic neutral loss of 91 Da is observed.The mechanism of the 91 Da loss was proposed and theoretically investigated.At the same time,the formation mechanism of the[c₁+2H]⁺fragment ion that appeared in the MS/MS spectrum was also discussed theoretically.The results achieved in this research are as below:1.By comparing multiple possible reaction pathways,peptide N-terminal radical induced the C_?–C_?bond cleavage mechanism refers to the optimal reaction pathway for the characteristic loss of 91 Da in the MS/MS spectrum of[FY]^·+.The barrier against this C_?–C_?bond cleavage and forming a hydrogen-bonded dimer is determined to be 25.4 kcal/mol(relative enthalpy at 0 K)and 24.8 kcal/mol(relative Gibbs free energy at 298 K).This dimer will then dissociate further into two product molecules with a relative enthalpy of 38.5kcal/mol(at 0 K)and relative Gibbs free energy of 24.9 kcal/mol(at 298 K).2.The formation mechanism of[c₁+2H]⁺fragment ion in the MS/MS spectrum of[FY]^·+was also discussed.The results showed that the formation of[c₁+2H]⁺fragment ion was related to the?-C radical on the tyrosine residue,which is generated through the proton transfer from this?-C atom to the nearby N atom.The barrier against this N–C_?bond cleavage and forming a hydrogen-bonded dimer is determined to be 29.1 kcal/mol(relative enthalpy at 0K)and 28.0 kcal/mol(relative Gibbs free energy at 298 K),which is slightly higher than the corresponding 91 Da loss pathway.This dimer will then dissociate further into two product molecules with a relative enthalpy of 28.9 kcal/mol(at 0 K)and relative Gibbs free energy of 14.1 kcal/mol(at 298 K).3.Based on the RRKM theory,the dynamic characteristics of 91 Da feature loss pathway and the[c₁+2H]⁺feature fragmentation pathway were analyzed.It is found that the rate constant of 91 Da feature loss dissociation pathway and[c₁+2H]⁺feature fragmentation have little difference under low[FY]^·+initial internal energy(<40 kcal/mol).As the increase of the internal energy,the dissociation rate constant of both pathways increase,and the increase of the reaction rate constants of the the 91Da feature loss pathway is more obvious.This conclusion is also in good agreement with the results of relevant experimental studies.It is hoped that our analysis of these characteristic fragment pathways can provide theoretical support for the identification of phenylalanine containing peptides.