Optical detection of cytochrome P450 by sensitizer-linked substrates

The ability to detect, characterize, and manipulate specific biomolecules i n complex media is critical for understanding metabolic processes. Particul arly important targets are oxygenases (cytochromes P450) involved in drug m etabolism and many disease states, including liver and kidney dysfunction, neurological disorders, and cancer. We have found that Ru photosensitizers linked to P450 substrates specifically recognize submicromolar cytochrome P 450(cam) in the presence of other heme proteins. In the P450:Ru-substrate c onjugates, energy transfer to the heme dramatically accelerates the Ru-lumi nescence decay, The crystal structure of a P450(cam):Ru-adamantyl complex r eveals access to the active center via a channel whose depth (Ru-Fe distanc e is 21 Angstrom) is virtually the same as that extracted from an analysis of the energy-transfer kinetics. Suitably constructed libraries of sensitiz er-linked substrates could be employed to probe the steric and electronic p roperties of buried active sites.