PROTEIN ENGINEERED HINGED GATE OPENS A CHANNEL TO AN ARTIFICIAL CAVITY. Duncan E. McRee*, Melissa M. Fitzgerald, Rabi A. Musah, and David B. Goodin, The Scripps Research Institute, 10666 N. Torrey Pines Rd., La Jolla, CA 92037

Conformational changes in the structures of proteins which gate the access of substrates or ligands to an active site are important features of enzyme function. We describe an unusual example of a structural rearrangement near a buried artificial cavity in cytochrome c peroxidase (Fitzgerald et al., 1994) upon binding of a positively charged benzimidazole that opens a channel to the buried cavity and apparently represents the entry of ligands to the buried cavity. A hinged rotation at two residues, Pro-190 and Asn-195, results in a surface loop rearrangement that opens a large solvent accessible channel to an otherwise inaccesible binding cavity. High resolution crystal structures have allowed detailed characterization of this rearrangement which includes a cis-tran proline isomerization and an asparagine swapping the side-chain oxygen for a main-chain carbonyl. In contrast, several structures of other bound ligands shows the gate to the cavity in a closed conformation and one ligand disorders the gate so that it is no longer visible in the electron denisty. Solution-state kinetics studies indicate that this conformational change is not an artifact of the crystallization conditions. The trapping of the open conformation provides a unique view of the extent to which protein dynamics can allow small molecule penetration into "inaccesible" protein cavities.

Fitzgerald, M.M., Churchill, M.J., McRee, D.E., and Goodin, D.B. (1995). "Small Molecule Binding to an Artificially Created Cavity at the Active Site of Cytochrome c Peroxidase." Biochemistry 33, 3807-3818.