Transient conformations in the unliganded FK506 binding domain of FKBP51 correspond to two distinct inhibitor-bound states.

Members of the FK506-binding protein (FKBP) family regulate a range of important physiological processes. Unfortunately, current therapeutics such as FK506 and rapamycin exhibit only modest selectivity among these functionally distinct proteins. Recent progress in developing selective inhibitors has been reported for FKBP51 and FKBP52 which act as mutual antagonists in the regulation of steroid hormone signaling. Two structurally similar inhibitors yield distinct protein conformations at the binding site. Localized conformational transition in the binding site of the unliganded FK1 domain of FKBP51 is suppressed by a K58T mutation that also suppresses the binding of these inhibitors. Here it is shown that the changes in amide hydrogen exchange kinetics arising from this K58T substitution are largely localized to this structural region. Accurate determination of the hydroxide-catalyzed exchange rate constants in both the wild-type and K58T variant proteins impose strong constraints upon the pattern of amide exchange reactivities within either a single or a pair of transient conformations that could give rise to the differences between these two sets of measured rate constants. Poisson-Boltzmann continuum dielectric calculations provide moderately accurate predictions of the structure-dependent hydrogen exchange reactivity for solvent exposed protein backbone amides. Applying such calculations to the local protein conformations observed in the two inhibitor-bound FKBP51 domains demonstrated that the experimentally determined exchange rate constants for the wild-type domain are robustly predicted by a population-weighted sum of the experimental hydrogen exchange reactivity of the K58T variant and the predicted exchange reactivities in model conformations derived from the two inhibitor-bound protein structures.