DEUTERIUM-EXCHANGE REACTIONS AS A PROBE OF BIOMOLECULE STRUCTURE - FUNDAMENTAL-STUDIES OF CAS PHASE H D EXCHANGE-REACTIONS OF PROTONATED GLYCINE OLIGOMERS WITH D2O, CD3OD, CD3CO2D, AND ND3/

A Fourier transform ion cyclotron resonance mass spectrometer was used to examine the hydrogen/deuterium exchange reactions of protonated gl ycine oligomers (Gly(n), n = 1-5) with D2O CD3OD, CD3CO2D, and ND3. Ex change rates in this study were monitored over three orders of magnitu de, from 10(-9) to 10(-12) cm(3) molecule(-1) s(-1). Reaction kinetics are highly dependent on peptide structure and the properties of the e xchange reagents. The rate and extent of H/D exchange of the protonate d oligomers increases with reagent gas basicity, D2O < CD3OD < CD3CO2D < ND3. ND3 is the most efficient reagent studied, as it exchanges eve ry labile hydrogen in each of the oligomers. Several distinct mechanis ms, supported by semiempirical AM1 and PM3 calculations, are proposed to explain the observed patterns of reactivity. An onium ion mechanism is proposed for the exchange of N-terminus hydrogens of Gly(n)H(+) ol igomers with ND3, in which an endothermic proton transfer from the N-t erminus is rendered energetically feasible by simultaneous solvation o f the resultant ammonium ion by the neutral peptide. This mechanism is consistent with the observation of multiple exchanges in a single col lision event with ND3. For those reagents whose proton affinities are too low to form solvated onium ion intermediates, a relay mechanism is proposed in which the reagent shuttles a proton from the N-terminus t o a slightly less basic site in the molecule. For glycine oligomers, t his sits is an amide oxygen. A tautomer mechanism is proposed for the exchange of the amide hydrogens with ND3. Exchange occurs by proton tr ansfer from the N-terminus to the amide carbonyl in concert with trans fer of the amide proton to ammonia, forming an ammonium ion solvated b y a tautomerized peptide. Semiempirical calculations suggest that exch ange of the C-terminus hydrogen proceeds via formtion of a salt bridge with the reagent gas, which deprotonates the C-terminus acid group, w ith the nearby protonated N-terminus stabilizing the resultant ion pai r. Betaine, [(CH3N+-CH2CO2H], used in this study to determine the isot opic purity of the exchange reagents, serves as a model for salt bridg e formation since it does not possess a labile proton and readily exch anges the carboxylic acid hydrogen. The effect of translational and vi brational excitation on H/D exchange rates was studied for several oli gomers using off-resonance collisional activation. For those oligomers that undergo facile H/D exchange with the reagent gases, excitation d ecreases rates. For those oligomers which do not undergo facile H/D ex change, reactivity is not promoted by collisional activation.