pKa Determination of Histidine Residues in α-Conotoxin MII Peptides by 1H NMR and Constant pH Molecular Dynamics Simulation
α-Conotoxin MII (α-CTxMII) is a potent and selective peptide antagonist of neuronal nicotinic acetylcholine receptors (nAChR’s). Studies have shown that His9 and His12 are signiﬁcant determinants of toxin binding aﬃnity for nAChR, while Glu11 may dictate diﬀerential toxin aﬃnity between nAChR isoforms. The protonation state of these histidine residues and therefore the charge on the α-CTx may contribute to the observed diﬀerences in binding aﬃnity and selectivity. In this study, we assess the pH dependence of the protonation state of His9 and His12 by 1H NMR spectroscopy and constant pH molecular dynamics (CpHMD) in α-CTxMII, α-CTxMII[E11A], and the triple mutant, α-CTxMII- [N5R:E11A:H12K]. The E11A mutation does not signiﬁcantly perturb the pKa of His9 or His12, while N5R:E11A:H12K results in a signiﬁcant decrease in the pKa value of His9. The pKa values predicted by CpHMD simulations are in good agreement with 1H NMR spectroscopy, with a mean absolute deviation from experiment of 0.3 pKa units. These results support the use of CpHMD as an eﬃcient and inexpensive predictive tool to determine pKa values and structural features of small peptides critical to their function.
McDougal, Owen M.; Granum, David M.; Swartz, Mark; Rohleder, Conrad; and Maupin, C. Mark. (2013). "pKa Determination of Histidine Residues in α-Conotoxin MII Peptides by 1H NMR and Constant pH Molecular Dynamics Simulation". Journal of Physical Chemistry B, 117(9), 2653-2661.