Peptides containing the chromophore, benzophenone (as the amino acid, 4-benzoyl-phenylalanine (Bpa)), have been synthesized to explore the feasibility of gas-phase photochemical cross-linking to investigate the conformations of unsolvated peptides. The main product of UV irradiation of a BP-containing peptide is CO2 loss from the peptide C terminus. To test whether decarboxylation results from hydrogen abstraction from acidic residues, we synthesized peptides designed to limit the contact between the Bpa and the peptide C terminus. Such contact is necessary for hydrogen abstraction. The peptide, Ac-Bpa-Ala(10)-Lys, forms an extended a-helix in the gas phase, thus preventing the N-terminal Bpa from approaching the C terminus.' In contrast, Ac-Bpa-Gly(10)-Lys is a flexible peptide that can adopt multiple "random globule" conformations in the gas phase, some of which could allow contact between the termini and thus photoinduced reaction. On simultaneous UV irradiation of a mixture of both electrosprayed gas-phase peptides in a Fourier transform ion cyclotron resonance ion trap, the degree of decarboxylation is 50-fold less for the a-helical alanine-based peptide than for the flexible glycine-based peptide. The results suggest that intimate contact between the photoexcited Bpa and the C terminus is necessary for decarboxylation, implicating hydrogen atom abstraction as the photochemical mechanism. These results establish the feasibility of probing gas-phase peptide/protein ion conformations by photochemistry of photoaffinity-labeled ions.