Probing Electron Transfer Processes in Biomacromolecules Using Transition Metal Complexes

摘要

Electron transfer (ET) reactions are crucial in numerous biological processes such as photosynthesis, cellular respiration, and DNA damage. Consequently, ET in biomacromolecules has been well-studied. Experiments using metals covalently attached to proteins have determined ET occurs by tunneling over short distances and hopping over long distances. Little work has been done to study ET in noncovalently bound systems. This thesis describes the use of biotinylated trinuclear ruthenium clusters as electrochemical probes for avidin binding. Using these clusters overcomes signal loss observed in previous experiments using smaller complexes. Binding constants measured by isothermal titration calorimetry for monovalent and bivalent biotinylated clusters were 1.2 - 8.0 x 106 M-. Modest change in electrochemical potential (-43 mV) was observed upon avidin binding for the bivalent system; however, no change in ET rate or reorganization energy was observed. Future work will tune ET parameters to maximize electrochemical changes upon protein binding.;Distance-dependent hopping and tunneling mechanisms have been established for oxidative hole transfer through DNA. Nevertheless, determination of the mechanism of ground&ndashstate ET remains elusive. This thesis describes the measurement of ground&ndashstate ET rates in DNA for elucidation of this mechanism. A flash&ndashquench scheme was used to measure the rate of ET between ruthenium donor and acceptor complexes across various lengths of DNA. A series of quenchers for use in this system was systematically studied. It was found that [Ru(NH3)6]3+ is the best quencher for the system. Additionally, the donor and acceptor ruthenium complexes were optimized. The long&ndashlived excited&ndashstate of the [Ru(bpy)2bpyTrzT]2+ acceptor resulted in greater formation of the mixed&ndashvalence metallated DNA species necessary for ET rate measurements. The optimized donor, [Ru(acac)2ImpyT] 0 has a much more facile synthesis than the previously used donor complex, [Ru(tolacac)2ImpyT]0. With this optimized flash-quench system, the rate of ET across metallated 10mer and 11mer oligonucleotides was found to be 3.8 x 106 s- and 1.3 x 106 s-, respectively. These rates suggest a tunneling mechanism; however, additional systems of different DNA lengths must be analyzed to convincingly establish the mechanism.