An electrospray ionization-flow tube study of H/D exchange in protonated bradykinin

An electrospray ionization-fast flow technique has. been employed to study the reactions of doubly protonated bradykinin and des-Arg(9)-bradykinin wit h CH3OD and ND3, respectively. Deconvolution of the experimental mass spect ral data followed by simulation of the kinetic data by solution of differen tial equations leads to sets of apparent and site specific rate constants. On a time scale of several milliseconds, bradykinin undergoes with ND3 thre e fast HID exchanges and one slow exchange. Three equivalent exchanges are observed with CH3OD that are nearly 2 orders of magnitude slower than the N D3 reactions. Up to six hydrogen exchanges are observed for the reaction of des-Arg(9)-bradykinin with ND3. The more efficient exchange of des-Arg(9)- bradykinin is accompanied by formation of collisionally stabilized complex es between doubly protonated des-Arg(9)-bradykinin and ND3 at a He carrier gas pressure of about 0.2 Torr. Multiple-collision activation-collision-ind uced dissociation of reactant and product ions of the isotope exchange reac tions was carried out in front of the sampling nose cone of the analyzer qu adrupole mass filter system. The degree of deuterium incorporation into the parent doubly protonated ions and into several of the b(n)(+) and y(n)(+) ions combined with the site-specific rate constants obtained indicates that the three equivalent hydrogens exchanged in doubly protonated bradykinin a re at the protonated N-terminus amine group. Complexation of doubly protona ted bradykinin by ND3 is prevented by its tightly folded structure, and thi s in turn prevents H/D exchange of the amide hydrogens of bradykinin. The a dditional H/D exchanges observed in the case of doubly protonated des-Arg(9 )-bradykinin are made possible by complexation of its less compact structur e via hydrogen-bonded intermediates that promote H/D exchange of amide hydr ogens.