title: Photoactive peptides for light-initiated tyrosyl radical generation and transport into ribonucleotide reductase.
abstract: The mechanism of radical transport in the alpha2 (R1) subunit of class I E. coli ribonucleotide reductase (RNR) has been investigated by the phototriggered generation of a tyrosyl radical, *Y356, on a 20-mer peptide bound to alpha2. This peptide, Y-R2C19, is identical to the C-terminal peptide tail of the beta2 (R2) subunit and is a known competitive inhibitor of binding of the native beta2 protein to alpha2. *Y356 radical initiation is prompted by excitation (lambda >or= 300 nm) of a proximal anthraquinone, Anq, or benzophenone, BPA, chromophore on the peptide. Transient absorption spectroscopy has been employed to kinetically characterize the radical-producing step by time resolving the semiquinone anion (Anq*-), ketyl radical (*-BPA), and Y* photoproducts on (i) BPA-Y and Anq-Y dipeptides and (ii) BPA/Anq-Y-R2C19 peptides. Light-initiated, single-turnover assays have been carried out with the peptide/alpha2 complex in the presence of [14C]-labeled cytidine 5'-diphosphate substrate and ATP allosteric effector. We show that both the Anq- and BPA-containing peptides are competent in deoxycytidine diphosphate formation and turnover occurs via Y731 to Y730 to C439 pathway-dependent radical transport in alpha2. Experiments with the Y730F mutant exclude a direct superexchange mechanism between C439 and Y731 and are consistent with a PCET model for radical transport in which there is a unidirectional transport of the electron and proton transport among residues of alpha2.
authors.affiliation: Department of Chemistry, Massachusetts Institute of Technology, 77 Massachusetts Avenue, Cambridge, Massachusetts 02139-4307, USA.
authors.forename: Steven Y
authors.initials: SY
authors.lastname: Reece
authors.forename: Mohammad R
authors.initials: MR
authors.lastname: Seyedsayamdost
authors.forename: Joanne
authors.initials: J
authors.lastname: Stubbe
authors.forename: Daniel G
authors.initials: DG
authors.lastname: Nocera
mesh_terms: D005609:Free Radicals; D008027:Light; D010455:Peptides / Q000737:chemistry*; D010777:Photochemistry; D012264:Ribonucleotide Reductases / Q000737:chemistry*; D014443:Tyrosine / Q000737:chemistry*
chemical_list: D005609:Free Radicals; D010455:Peptides; D014443:Tyrosine; D012264:Ribonucleotide Reductases