ABSTRACT
Escherichia coli ribonucleotide reductase (RNR) catalyzes the conversion of nucleoside diphosphates (NDPs) to deoxynucleoside diphosphates (dNDPs). This RNR is composed of two homodimeric subunits: R1 and R2. R1 binds the NDPs in the active site, and R2 harbors the essential di-iron tyrosyl radical (Y*) cofactor. In this paper, we used PELDOR, a method that detects weak electron-electron dipolar coupling, to make the first direct measurement of the distance between the two Y*'s on each monomer of R2. In the crystal structure of R2, the Y*'s are reduced to tyrosines, and consequently R2 is inactive. In R2, where the Y*'s assume a well-defined geometry with respect to the protein backbone, the PELDOR method allows measurement of a distance of 33.1 +/- 0.2 A that compares favorably to the distance (32.4 A) between the center of mass of the spin density distribution of each Y* on each R2 monomer from the structure. The experiments provide the first direct experimental evidence for two Y*'s in a single R2 in solution.
Subject(s)
Escherichia coli/enzymology , Ribonucleotide Reductases/chemistry , Tyrosine/chemistry , Computer Simulation , Electron Spin Resonance Spectroscopy/methods , Free Radicals/chemistryABSTRACT
Here, we report a novel strategy for the combinatorial or parallel solid-phase synthesis of potential inhibitors of the mur-pathway enzymes. The strategy involves the efficient use of p-alkoxybenzylidene acetal linker for reversible immobilization of sugar scaffolds to solid phase. This methodology was used to synthesize several glycopeptides on solid phase in good yields.