Competition between diketopiperazine and oxazolone formation in water loss products from protonated ArgGly and GlyArg

The mechanism of peptide “b” fragment formation in collision-induced dissociation (CID) is generally understood as a nucleophilic attack from a carbonyl oxygen onto the electron deficient carbon of the dissociating amide bond forming a five-membered oxazolone ring structure. Nonetheless, other nucleophiles, such as the N-terminus and side-chain moieties (e.g., imidazole, guanidine), can in principle engage in a nucleophilic attack to induce amide backbone cleavage. Here, we apply a combination of infrared multiple photon dissociation (IRMPD) spectroscopy and computational chemistry to characterize the water loss, [M+H-H2O]+, product ions from protonated ArgGly and GlyArg. IRMPD spectra for [M+H-H2O]+ from ArgGly and GlyArg differ in the presence and absence of a characteristic band at 1885 cm−1, which is indicative of an oxazolone structure for ArgGly. The remaining parts of the vibrational spectra are consistent with the vibrational signatures of diketopiperazine structures. Conversely, there is no match between the experimental spectra and any of the putative structures arising from guanidine side-chain attack. These results show that the presence of a basic residue, such as arginine, facilitates the formation of diketopiperazine structures, and that residue order matters in the competition between diketopiperazine and oxazolone pathways.

Graphical abstractFigure optionsDownload full-size imageDownload high-quality image (112 K)Download as PowerPoint slideHighlights► The water loss products from protonated ArgGly and GlyArg are investigated by infrared multiple photon dissociation spectroscopy and computational chemistry. ► While ArgGly exclusively yields diketopiperazine structures, a mixture of diketopiperazine and oxazolone structures are confirmed for GlyArg. ► Product structures due to nucleophilic attacks from the guanidine side chain can be excluded.