Leaving group and gas phase neighboring group effects in the side chain losses from protonated serine and its derivatives

The gas phase fragmentation reactions of protonated serine and its YNHCH(CH₂X)CO₂H derivatives, β-chloroalanine, S-methyl cysteine, O-methyl serine, and O-phosphoserine, as well as the corresponding N-acetyl model peptides have been examined via electrospray ionization tandem mass spectrometry (MS/MS). In particular, the competition between losses from the side chain and the combined loss of H₂O and CO from the C-terminal carboxyl group of the amino acids or H₂O or CH₂CO from the N-acetyl model peptides are compared. In this manner the effect of the leaving group (Y = H or CH₃CO, vary X) or of the neighboring group can be examined. It was found that the amount of HX lost from the side chain increases with the proton affinity of X OP(O)(OH)₂ > OCH₃ ≈ OH > Cl]. The ion due to the side chain loss of H₂O from the model peptide N-acetyl serine is more abundant than that from protonated serine, suggesting that the N-acetyl group is a better neighboring group than the amino group. Ab initio calculations at the MP2(FC)/6-31G*//HF/6-31G* level of theory suggest that this effect is due to the transition state barrier for water loss from protonated N-acetyl serine being lower than that for protonated serine. The mechanism for side chain loss has been examined using MS³ tandem mass spectrometry, independent synthesis of proposed product ion structures combined with MS/MS, and hydrogen/deuterium exchange. Neighboring group rather than cis 1,2 elimination processes dominate in all cases. In particular, the loss of H₃PO₄ from O-phosphoserine and N-acetyl O-phosphoserine is shown to yield a 3-membered aziridine ring and 5-membered oxazoline ring, respectively, and not the dehydroalanine moiety. This is in contrast to results presented by DeGnore and Qin (J. Am. Soc. Mass Spectrom. 1998, 9, 1175–1188) for the loss of H₃PO₄ from larger peptides, where dehydroalanine was observed. Alternate mechanisms to cis 1,2 elimination, for the formation of dehydroalanine in larger phosphoserine or phosphothreonine containing peptides, are proposed.