UC Davis

Putidaredoxin (Pdx) is the exclusive reductase and a structural effector for P450cam (CYP101A1). However, the mechanism of how Pdx modulates the conformational states of P450cam remains elusive. Here we report a putative communication pathway for the Pdx-induced conformational change in P450cam using results of double electron-electron resonance (DEER) spectroscopy and molecular dynamics simulations. Use of solution state DEER measurements allows us to observe subtle conformational changes in the internal helices in P450cam among closed, open, and P450cam-Pdx complex states. Molecular dynamics simulations and dynamic network analysis suggest that Pdx binding is coupled to small coordinated movements of several regions of P450cam, including helices C, B', I, G, and F. These changes provide a linkage between the Pdx binding site on the proximal side of the enzyme and helices F/G on the distal side and the site of the largest movement resulting from the Pdx-induced closed-to-open transition. This study provides a detailed rationale for how Pdx exerts its long-recognized effector function at the active site from its binding site on the opposite face of the enzyme.