Gas-phase collision induced dissociation mechanisms of peptides: Theoretical and experimental study of N-formylalanylamide fragmentation

In order to shed light on the fragmentation mechanisms occurring during the collision induced dissociation (CID) of peptides in the gas phase, we have studied a model system, the N-formylalanylamide (HCO-Ala-NH2), by coupling experimental and theoretical methods. In particular, we have addressed two different questions arising in such experiments: (i) what is (are) the structure(s) of the ion before collision, and (ii) what are the fragmentation mechanisms occurring after collision with the target gas. For the first question, we coupled the potential energy surface (PES) study done by means of density functional theory (DFT), with Infra Red Multiple Photon Dissociation (IRMPD) spectroscopy. For the second problem, which is actually the main topic of the present work, we coupled quantum mechanics plus molecular mechanics (QM + MM) direct chemical dynamics simulations with tandem mass spectrometry (MS/MS). In addition, in order to better delineate the fragmentation mechanisms and validate those proposed by simulations, isotopic labeling experiments using 2H and 13C were performed. Thanks to the interplay between simulations and experiments, it was possible to successfully identify the fragmentation pathways leading to b1, y1, a1 and immonium ions. Our mechanisms support the “mobile proton” picture that is supposed to trigger the peptide fragmentation in the gas phase, confirming, from a chemical dynamics point of view, previous theoretical and experimental studies on similar systems.

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► Collision induced dissociation mechanisms of a simple modified amino acid, the N-formylalanylamide (HCO-Ala-NH2), were studied by QM + MM chemical dynamics simulations and experiments.
► IRMPD experiments and DFT calculations were performed to identify the species present in the gas phase before collision.
► HCO-Ala-NH2 and the labeled DCO-Ala-NH2 and H13CO-Ala-NH2 have been synthesized for the first time and CID experiments done to confirm the reaction mechanisms.
► The experimental MS/MS fragments were successfully identified from simulations and the proposed mechanisms are confirmed by the 2H and 13C labeled experiments.
► We have observed that also direct dynamics is in agreement with the “mobile proton” model to better understand MS/MS fragmentation.