title: Generation of a stable, aminotyrosyl radical-induced α2β2 complex of Escherichia coli class Ia ribonucleotide reductase.
abstract: Ribonucleotide reductase (RNR) catalyzes the conversion of nucleoside diphosphates to deoxynucleoside diphosphates (dNDPs). The Escherichia coli class Ia RNR uses a mechanism of radical propagation by which a cysteine in the active site of the RNR large (α2) subunit is transiently oxidized by a stable tyrosyl radical (Y•) in the RNR small (β2) subunit over a 35-Å pathway of redox-active amino acids: Y122• ↔ [W48?] ↔ Y356 in β2 to Y731 ↔ Y730 ↔ C439 in α2. When 3-aminotyrosine (NH2Y) is incorporated in place of Y730, a long-lived NH2Y730• is generated in α2 in the presence of wild-type (wt)-β2, substrate, and effector. This radical intermediate is chemically and kinetically competent to generate dNDPs. Herein, evidence is presented that NH2Y730• induces formation of a kinetically stable α2β2 complex. Under conditions that generate NH2Y730•, binding between Y730NH2Y-α2 and wt-β2 is 25-fold tighter (Kd = 7 nM) than for wt-α2
authors: Department of Chemistry, Massachusetts Institute of Technology, Cambridge, MA 02139, USA.
authors: Ellen C
authors: EC
authors: Minnihan
authors: Nozomi
authors: N
authors: Ando
authors: Edward J
authors: EJ
authors: Brignole
authors: Lisa
authors: L
authors: Olshansky
authors: Johnathan
authors: J
authors: Chittuluru
authors: Francisco J
authors: FJ
authors: Asturias
authors: Catherine L
authors: CL
authors: Drennan
authors: Daniel G
authors: DG
authors: Nocera
authors: Joanne
authors: J
authors: Stubbe
mesh_terms: D020134:Catalytic Domain; D004579:Electron Transport; D004795:Enzyme Stability; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics; D029968:Escherichia coli Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D007700:Kinetics; D008854:Microscopy, Electron; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed; D020836:Protein Structure, Quaternary; D021122:Protein Subunits; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012264:Ribonucleotide Reductases / Q000737:chemistry* / Q000145:classification / Q000235:genetics / Q000378:metabolism*; D053838:Scattering, Small Angle; D013050:Spectrometry, Fluorescence; D014961:X-Ray Diffraction
chemical_list: D029968:Escherichia coli Proteins; D021122:Protein Subunits; D011994:Recombinant Proteins; D012264:Ribonucleotide Reductases