Re(bpy)(CO)[subscript 3]CN as a Probe of Conformational Flexibility in a Photochemical Ribonucleotide Reductase
Author(s)
Lutterman, Daniel; Seyedsayamdost, Mohammad R.; Stubbe, JoAnne; Reece, Steven Y., 1980-; Nocera, Daniel G., 1957-
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Photochemical ribonucleotide reductases (photoRNRs) have been developed to study the proton-coupled electron transfer (PCET) mechanism of radical transport in Escherichia coli class I ribonucleotide reductase (RNR). The transport of the effective radical occurs along several conserved aromatic residues across two subunits: β2([superscript •]Y122 → W48 → Y356) → α2(Y731 → Y730 → C439). The current model for RNR activity suggests that radical transport is strongly controlled by conformational gating. The C-terminal tail peptide (Y-βC19) of β2 is the binding determinant of β2 to α2 and contains the redox active Y356 residue. A photoRNR has been generated synthetically by appending a Re(bpy)(CO)[subscript 3]CN ([Re]) photo-oxidant next to Y356 of the 20-mer peptide. Emission from the [Re] center dramatically increases upon peptide binding, serving as a probe for conformational dynamics and the protonation state of Y356. The diffusion coefficient of [Re]-Y-βC19 has been measured (k[subscript d1] = 6.1 × 10[superscript −7] cm[superscript −1] s[superscript −1]), along with the dissociation rate constant for the [Re]-Y-βC19−α2 complex (7000 s[superscript −1] > k[subscript off] > 400 s[superscript −1]). Results from detailed time-resolved emission and absorption spectroscopy reveal biexponential kinetics, suggesting a large degree of conformational flexibility in the [Re]-Y-βC19−α2 complex that engenders partitioning of the N-terminus of the peptide into both bound and solvent-exposed fractions.
Date issued
2009-04Department
Massachusetts Institute of Technology. Department of Biology; Massachusetts Institute of Technology. Department of ChemistryJournal
Biochemistry
Publisher
American Chemical Society (ACS)
Citation
Reece, Steven Y., Daniel A. Lutterman, Mohammad R. Seyedsayamdost, JoAnne Stubbe, and Daniel G. Nocera. “Re(bpy)(CO)3CN as a Probe of Conformational Flexibility in a Photochemical Ribonucleotide Reductase.” Biochemistry 48, no. 25 (June 30, 2009): 5832-5838.
Version: Author's final manuscript
ISSN
0006-2960
1520-4995