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Conformationally Regulated Peptide Bond Cleavage in Bradykinin

Fuller, Daniel R., Conant, Christopher R., El-Baba, Tarick J., Brown, Christopher J., Woodall, Daniel W., Russell, David H., Clemmer, David E.
Journal of the American Chemical Society 2018 v.140 no.30 pp. 9357-9360
biological resistance, bradykinin, cleavage (chemistry), humans, isomerization, mass spectrometry, temperature
Ion mobility and mass spectrometry techniques are used to investigate the stabilities of different conformations of bradykinin (BK, Arg¹-Pro²-Pro³-Gly⁴-Phe⁵-Ser⁶-Pro⁷-Phe⁸-Arg⁹). At elevated solution temperatures, we observe a slow protonation reaction, i.e., [BK+2H]²⁺+H⁺ → [BK+3H]³⁺, that is regulated by trans → cis isomerization of Arg¹-Pro², resulting in the Arg¹-cis-Pro²-cis-Pro³-Gly⁴-Phe⁵-Ser⁶-cis-Pro⁷-Phe⁸-Arg⁹ (all-cis) configuration. Once formed, the all-cis [BK+3H]³⁺ spontaneously cleaves the bond between Pro²-Pro³ with perfect specificity, a bond that is biologically resistant to cleavage by any human enzyme. Temperature-dependent kinetics studies reveal details about the intrinsic peptide processing mechanism. We propose that nonenzymatic cleavage at Pro²-Pro³ occurs through multiple intermediates and is regulated by trans → cis isomerization of Arg¹-Pro². From this mechanism, we can extract transition state thermochemistry: ΔG‡ = 94.8 ± 0.2 kJ·mol–¹, ΔH‡ = 79.8 ± 0.2 kJ·mol–¹, and ΔS‡ = −50.4 ± 1.7 J·mol–¹·K–¹ for the trans → cis protonation event; and, ΔG‡ = 94.1 ± 9.2 kJ·mol–¹, ΔH‡ = 107.3 ± 9.2 kJ·mol–¹, and ΔS‡ = 44.4 ± 5.1 J·mol–¹·K–¹ for bond cleavage. Biological resistance to the most favored intrinsic processing pathway prevents formation of Pro³-Gly⁴-Phe⁵-Ser⁶-cis-Pro⁷-Phe⁸-Arg⁹ that is approximately an order of magnitude more antigenic than BK.