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Design, characterization, and electron transfer properties of synthetic metalloproteins

by Hong, Jing.

Abstract (Summary)
Michael Ogawa, Advisor The binding of Cu(I) to the random coil peptide C16C19-GGY produces a self-organized metal-peptide assembly which displays an intense room-temperature luminescence at 600 nm. It was shown that this synthetic metalloprotein exists as a 4-helix bundle which contains a cyclic Cu4S4 cofactor in which each Cu(I) atom is bridged by two cysteine residues and has a terminal N/O ligand. The strong luminescence of the Cu(I) protein suggests that it might function as a photoinduced electron-transfer agent. The emission follows biexponential decay kinetics with ?1 = 1.0 µsec and ?2 = 7.5 µsec. These components have approximately equal amplitudes and the results indicate that the Cu4S4 cofactor contains two independent lumophores. Both lifetime components are quenched by a series of [Ru(NH3)5L]3+ (L = chloro, amine, lutidine, pyridine, nicotinamide, and 3,5-dimethyl pyridine dicarboxylate). The quenching mechanism is assigned to a photoinduced electron-transfer event by transient spectroscopy. The results show the occurrence of bimolecular forward electron-transfer in the inverted Marcus regime. Electron-transfer (ET) reactions occur between a negatively charged cyclic metallopeptide [Ru(bpy)2(phen-am)-cyclo(Cys-Glu-D-Glu-Glu-Pro-Glu-D-Glu)]3- = Rucyclic, and ferricytochrome c = cyt c, in which an acetamido linker was used to attach the ruthenium polypyridyl complex to the cysteine side chain of a head to tail cyclic peptide. In the presence of cyt c, the triplet state of ruthenium metallopeptide decays via parallel pathways that involve two different encounter peptide-protein complexes. That the electron transfer rate constants of both encounter complexes decrease with increasing viscosity demonstrates that the kinetics are gated by rate-limiting configurational changes occurring within the complexes. NMR experiments iii confirm that two separate conformations exist for Rucyclic. The two conformations of Rucyclic might be arising from the cis/trans isomerization of proline and result in the formation of two encounter complexes when interact with cyt c. Metallopeptides, 5-Chloro-PhenRuCE5G with a different redox potential but a similar conformation with those of RuCE5G give different driving forces of the excited-state ET reaction. The reorganization energy (?) and donor-acceptor separation (r) of the preformed complex between metallopeptides and cyt c are determined to be 1.25 eV and 16.4 Å by measuring the actual electron transfer rates of these four metallopeptides at their most stable configurations. These values are comparable with those of some protein-protein systems reported previously. iv To my parents, Fangzheng Hong and Guilan Liu, and to my sister Lin Hong for their love and support. v
Bibliographical Information:

Advisor:

School:Bowling Green State University

School Location:USA - Ohio

Source Type:Master's Thesis

Keywords:metalloproteins charge exchange

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