SUBSTRATE MOBILITY IN THIOCAMPHOR-BOUND CYTOCHROME-P450(CAM) - AN EXPLANATION OF THE CONFLICT BETWEEN THE OBSERVED PRODUCT PROFILE AND THE X-RAY STRUCTURE

Thiocamphor is an unusual substrate for P450cam in that in the X-ray s tructure it binds in the active site pocket in two distinct orientatio ns and neither of these orientations are consistent with the 5-alcohol being the primary product. Other camphor analogs such as norcamphor o r camphane bind in a single orientation consistent with the 5-alcohol being a major product. We present an analysis of four 175 ps molecular dynamics trajectories of thiocamphor-bound cytochrome P450cam. The fi rst two trajectories were calculated for cytochrome P450cam with thioc amphor bound in both its major and minor crystallographic orientations . In the second set of simulations, a single oxygen atom was added as a distal ligand to the heme group in order to model the putative ferry l oxygen reaction intermediate. Trajectories were again calculated sta rting with thiocamphor in its major and minor orientations. While the protein dynamics were quite similar in atl four trajectories, the subs trate showed distinctly different motions in each of the trajectories. In particular, the preferred substrate orientations were very differe nt in the presence of the ferryl oxygen than in the absence of that ox ygen. The preferred orientations in the absence of the distal oxygen w ere consistent with the 3-alcohol being the major product, while the p referred orientations in the presence of the distal oxygen were consis tent with the 5-alcohol being a major product. These simulations offer an explanation for the inconsistency between the X-ray data and the p roduct profile.