title: Function of the diiron cluster of Escherichia coli class Ia ribonucleotide reductase in proton-coupled electron transfer.
abstract: The class Ia ribonucleotide reductase (RNR) from Escherichia coli employs a free-radical mechanism, which involves bidirectional translocation of a radical equivalent or "hole" over a distance of ~35 Å from the stable diferric/tyrosyl-radical (Y122(•)) cofactor in the β subunit to cysteine 439 (C439) in the active site of the α subunit. This long-range, intersubunit electron transfer occurs by a multistep "hopping" mechanism via formation of transient amino acid radicals along a specific pathway and is thought to be conformationally gated and coupled to local proton transfers. Whereas constituent amino acids of the hopping pathway have been identified, details of the proton-transfer steps and conformational gating within the β sununit have remained obscure; specific proton couples have been proposed, but no direct evidence has been provided. In the key first step, the reduction of Y122(•) by the first residue in the hopping pathway, a water ligand to Fe1 of the diferric cluster was suggested to donate a proton to yield the neutral Y122. Here we show that forward radical translocation is associated with perturbation of the Mössbauer spectrum of the diferric cluster, especially the quadrupole doublet associated with Fe1. Density functional theory (DFT) calculations verify the consistency of the experimentally observed perturbation with that expected for deprotonation of the Fe1-coordinated water ligand. The results thus provide the first evidence that the diiron cluster of this prototypical class Ia RNR functions not only in its well-known role as generator of the enzyme's essential Y122(•), but also directly in catalysis.
authors.affiliation: Department of Chemistry, The Pennsylvania State University, University Park, Pennsylvania 16802, USA.
authors.forename: Bigna
authors.initials: B
authors.lastname: Wörsdörfer
authors.forename: Denise A
authors.initials: DA
authors.lastname: Conner
authors.forename: Kenichi
authors.initials: K
authors.lastname: Yokoyama
authors.forename: Jovan
authors.initials: J
authors.lastname: Livada
authors.forename: Mohammad
authors.initials: M
authors.lastname: Seyedsayamdost
authors.forename: Wei
authors.initials: W
authors.lastname: Jiang
authors.forename: Alexey
authors.initials: A
authors.lastname: Silakov
authors.forename: JoAnne
authors.initials: J
authors.lastname: Stubbe
authors.forename: J Martin
authors.initials: JM
authors.lastname: Bollinger
authors.forename: Carsten
authors.initials: C
authors.lastname: Krebs
mesh_terms: D004579:Electron Transport; D004926:Escherichia coli / Q000201:enzymology* / Q000378:metabolism; D005290:Ferric Compounds / Q000737:chemistry / Q000378:metabolism*; D015394:Molecular Structure; D011522:Protons; D012264:Ribonucleotide Reductases / Q000737:chemistry / Q000145:classification / Q000378:metabolism*
chemical_list: D005290:Ferric Compounds; D011522:Protons; D012264:Ribonucleotide Reductases