Griselda Hernández, Ph.D.

Using NMR spectroscopy and molecular design, we characterize the coupled transitions of FK506-binding domain proteins to help understand their roles in regulatory signaling and to facilitate selective structure-based therapeutic design. We have primarily applied these approaches to two pairs of closely homologous human FKBP proteins: FKBP12 and FKBP12.6, which regulate the ryanodine receptor calcium channels of skeletal and cardiac muscle, and the first FKBP domains of FKBP51and FKBP52, best characterized for their antagonistic roles in regulating the transcriptional activity of various steroid hormone receptor complexes. Reflecting an increased research interest in using FK506 and related therapeutics in treating pathogenic fungal infections, we are applying such structural dynamics analyses to the FKBP12 protein of highly lethal Candida auris which the Centers of Disease Center recently designated as the first fungal pathogen to be listed among the five most urgent threats in their Antibiotic Resistance Threats Report.

Much of the challenge from this line of therapeutic development stems from the fact that the parent drug FK506 and the closely related rapamycin strongly inhibit most members of the much larger FKBP family of proteins. Furthermore, both FK506 and rapamycin are primarily used clinically for their immunosuppressive effects following tissue transplantation surgery. While earlier drug design studies have demonstrated reduction in the immunosuppressive effects of FK506-related compounds, there still remains the need to develop clinical selectivity among the individual members of the FKBP family which has been confounded by the close similarities among the drug binding sites seen in the crystal structures of the various FKBP proteins.

In contrast, our NMR relaxation studies of the human FKBP binding domains have demonstrated markedly differing patterns of conformational dynamics among FKBP12, FKBP12.6, FKBP51, and FKBP52 which reflect differences in the structural transitions that each of these domains undergo. In particular, we have demonstrated that one such conformational transition that is observed only in the NMR spectra of FKBP51 domain which corresponds closely to the structural reorganization more recently reported in the crystal structure of the first FKBP51-selective inhibitor. This localized transition is, in turn, energetically coupled to a second distinct conformational transition of FKBP51 involving the site of interaction with the steroid receptor protein. Neither transition is observed in solution for the FKBP52 domain. Mutations that interchange the inhibitory (FKBP51) and stimulatory (FKBP52) behavior of these two proteins also interchange the dynamical properties in this region of the protein. We have also found that, among these four FKBP domains, only FKBP12 exhibits a characteristic conformational transition within the drug-binding site which might prove amenable to drug design engineering.