The iron center in ribonucleotide reductase from Escherichia coli.

Abstract

Ribonucleotide reductase from Escherichia coli consists of two nonidentical subunits, proteins B1 and B2. The active site is made up from both subunits. Protein B2 contains 2 iron atoms and a tyrosyl-free radical, which are essential for the enzymatic activity. The paramagnetic susceptibility of protein B2 has been measured over the temperature range 30-200 K. A deviation from the Curie law is observed at high temperatures, consistent with a structure of an antiferromagnetically coupled pair of high spin Fe(III) with an exchange coupling -J = 108(-20)+25 cm-1. Electronic spectra are resolved into components from the iron center and the radical. A band at 600 nm is clearly identified and shown to have contributions from both components. The electronic absorptions of the tyrosyl radical of protein B2 are closely similar to those reported for phenoxy radicals of tyrosine and tritertiary butyl phenol. Determinations by EPR of the amount of free radical suggest the possibility of more than one radical per active protein B2 molecule. Reconstitution of the active site from apoprotein B2 and Fe(II) is only observed in the presence of oxygen. With Fe(III), no reconstitution is obtained. The additional physical data on the iron center of protein B2 strengthen the analogy with oxidized forms of hemerythrin. The most likely structure is an antiferromagnetically coupled pair of high spin Fe(III), possibly with a bridging oxo-group.