Diiron carboxylate proteins contain a cofactor that consists of two iron atoms coordinated by carboxylate and histidine ligands. These proteins perform a multitude of chemical reactions in the cell that generally involve activation of molecular oxygen at the diiron site.
Ribonucleotide reductase is the only enzyme that performs de novo synthesis of all four deoxyribonucleotides, the building blocks of DNA. The R2 protein of Class-I ribonucleotide reductase, which is a diiron carboxylate protein, utilises the high-valent iron-oxygen species generated at the diiron site to produce a stable tyrosyl radical required for enzymatic activity.
In this work, X-ray crystallography and EPR are used to study the R2 protein from Escherichia coli with the goal of understanding its mechanism of oxygen activation, radical generation and radical stabilisation. Based on these studies a detailed structural mechanism is proposed, which might be common to several oxygen activating diiron carboxylate proteins. The orientation of the active radical species in R2 has also been determined. In addition, these results provide a rationale for the unusual stability of the radical.
Crystal structures of R2 proteins from two other species, Corynebacterium ammoniagenes and Chlamydia trachomatis, have also been solved.
The C. ammoniagenes protein has been reported to be manganese-dependent. The results presented here, however, support dependence of iron and not manganese.
Sequence alignments indicate that the chlamydial R2s lack the, otherwise conserved, radical harbouring tyrosine. This is confirmed by the structure, which also reveals other unique features in the diiron site. Hypotheses regarding the function of the protein and the reason for the differences are presented.
2003. , 77 p.