Re(bpy)(CO)[subscript 3]CN as a Probe of Conformational Flexibility in a Photochemical Ribonucleotide Reductase
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Author(s)
Lutterman, Daniel
Seyedsayamdost, Mohammad R.
Stubbe, JoAnne
Reece, Steven Y., 1980-
Nocera, Daniel G., 1957-
Date Issued
April 2009
Journal
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.
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Author's final manuscript
Abstract
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.
MIT Department
Massachusetts Institute of Technology. Department of Biology
Massachusetts Institute of Technology. Department of Chemistry
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DOI of Published Version
https://doi.org/10.1021/bi9005804
