TABLE 3

Allosteric parameter estimates for gallamine, strychnine, and brucine at the WT and N410K+T423K M2 mAChRs

Values represent the mean ± S.E. from at least three experiments performed in duplicate.

WTN410K+T423K
pKBaLogαNMSbnckoff (min−1)dncpKBLogαNMSnkoff (min−1)n
Gallamine5.98 ± 0.05−1.15 ± 0.0560.0056 ± 0.0003^43.91 ± 0.20*−0.57 ± 0.1130.010 ± 0.0023
Strychnine5.48 ± 0.300.38 ± 0.0630.0009 ± 0.0002^3n.an.a40.004 ± 0.0013
Brucine5.58 ± 0.340.14 ± 0.0330.0014 ± 0.0003^34.34 ± 0.34*−0.36 ± 0.1840.002 ± 0.0013
  • n.a., not applicable, as no alteration of the specific binding of the radioligand was observed, and therefore, no estimate could be provided.

  • a Negative logarithm of the allosteric modulator equilibrium dissociation constant.

  • b Logarithm of the cooperativity factor for the interaction between the modulators and the orthosteric radioligand.

  • c Number of experiments performed.

  • d Dissociation rate constant of the [3H]NMS in the presence of the indicated ligand (control values in the absence of ligand: WT koff = 0.043 ± 0.003 min−1, N410K+T423K mutant koff = 0.007 ± 0.001 min−1).

  • * Significantly different from the corresponding value at the wild type (P < 0.05), Student’s t test.

  • ^ Significantly different from koff with 100 μM atropine only (P < 0.05), one-way analysis of variance with Dunnett’s post-test.