General Colloquium
April 19 - 4:00pm Phys 223
(Coffee at 3:30p.m. in room 242)
April 19 - 4:00pm Phys 223
(Coffee at 3:30p.m. in room 242)
Stephen G. Sligar
Professor Chemistry, Biochemistry and the College of Medicine
University of Illinois, Beckman Institute, Urbana, Illinois 61801
Title:"Cryocrystallography and Cryoenzymology of the Cytochromes P450"
Members of the cytochrome P450 superfamily catalyze the addition of molecular oxygen to non-activated hydrocarbons at physiological temperature - a reaction that normally requires high temperature to proceed in the absence of a catalyst. Working in collaboration with professors Petsko, Schlichting, Hoffman, and Kincaid together with their coworkers, structures were obtained for the significant intermediates in the hydroxylation reaction of a P450 monoxygenase using trapping techniques and cryo-crystallography. The structure of the ferrous dioxygen adduct of P450cam was determined using 0.91 Å wavelength X-rays, a wavelength that minimizes radiation chemistry produced redox equivalents. Irradiation with 1.5 Å X-rays results in cleavage of the bound dioxygen in the crystal. High resolution structures show the conformational changes of several important residues which are linked to a network of bound water molecules that provide the protons needed for this reaction We have also been able to directly follow the sequence of catalytic events in solution by reacting the ferrous dioxygen intermediate with radiolytically produced reducing equivalents from in situ phosphorous-32 or exogenous cobalt-60 and following the heme and proton states using EPR, ENDOR, Raman and optical spectroscopy. With step-wise thermal annealing, these techniques identify sequential iron-oxygen intermediates and offer an integrated picture of Cytochrome P450 monoxygenase catalysis.