This proposal seeks to investigate the mechanisms of biological electron-transfer (ET) reactions through the synthesis and study of de novo metalloproteins that have the native-like structure of an alpha-helical coiled-coil. An important feature of these peptides is that they provide the first well-defined model system in which to study the mechanisms of biological ET reactions that occur across a non-covalent peptide-peptide interface. They can also be derivatized to study ET reactions that occur along the covalent backbone of an alpha-helix. A 30-mer polypeptide has recently been prepared and shown to self-assemble into a non-covalent, two-stranded alpha-helical coiled-coil. Metal-based redox centers (i.e. Ru(bpy)(trpy)Im-, and Ru(NH3)5-) were coordinated to histidine residues located at equivalent positions on each of the two peptide chains to prepare the electron-transfer metalloprotein which has been characterized by a number of analytical methods. Pulse radiolysis was used to provide the first observation of an electron-transfer reaction occurring across a non-covalent peptide-peptide interface in a model protein with k(et) = 9600 +- 700 s-1. The experiments described in this proposal will further examine the ET properties of alpha-helical coiled-coils. Studies will be conducted to help determine how, or if, various chemical alterations of the non-covalent peptide interface can influence the rate of electron-transfer. Studies will also be conducted on reactions that occur across a covalently crosslinked interface. Finally, ET reactions will be studied that proceed along the covalent backbone of a single alpha-helical peptide chain. Future results from this work should provide valuable information about the nature of protein-based electron-transfer processes.
