PMID:10830848
| Citation |
Twitchett, MB, Dobbing, AM and Sykes, AG (2000) New mechanistic insights into the reactivity of the R2 protein of E. coli ribonucleotide reductase (RNR). J. Inorg. Biochem. 79:59-65 |
|---|---|
| Abstract |
Further to a linear free-energy correlation of cross-reaction rate constants k12 for the reaction of eight organic radicals (OR), e.g. MV*+, from methyl viologen, with cytochrome c(III), we consider here similar studies for the reduction of the R2 protein of Escherichia coli ribonucleotide reductase, which has FeIII2 and Tyr* redox components. The same two techniques of pulse radiolysis and stopped-flow were used. Cross-reaction rate constants (22 degrees C) at pH 7.0, I=0.100 M (NaCl), were determined for the reduction of active-R2 with the eight ORs, reduction potentials E0(1) from -0.446 to +0.194 V. Samples of active-R2 have an FeIII2 met-R2 component, which in the present studies was close to 40%. Concurrent reactions have to be taken into account for the five most reactive ORs, corresponding to reduction of the FeIII2 of met-R2 and then of active-R2. Separate experiments on met-R2 reproduced the first of these rate constants, which on average is approximately 66% larger than the second rate constant. A single Marcus free-energy plot of log k12-0.5 log10f versus -E0(1)/0.059 describes all the data and the slope of 0.54 is in satisfactory agreement with the theoretical value of 0.50. Such behaviour is unexpected since the Tyr* is a much stronger oxidant (E0 approximately 1.0 V versus NHE) as compared to FeIII2 (E0 close to zero). X-ray structures of the met- and red-R2 states have indicated that electroneutrality of the approximately 10 A buried active site is maintained. Proton transfer is therefore proposed as a rapid sequel to electron transfer. Other reactions considered are the much slower conventional time-range reductions of active-R2 with hydrazine and dithionite. For these reactions one and/or two-equivalent changes are possible. With both reductants, met-R2 reacts about four-fold faster than active-R2, and as with the ORs the less strongly oxidising FeIII2 component is reduced before the Tyr*. |
| Links | |
| Keywords |
Buffers; Escherichia coli/enzymology; Hydrogen-Ion Concentration; Indicators and Reagents; Iron/metabolism; Kinetics; Models, Chemical; Oxidation-Reduction; Ribonucleotide Reductases/chemistry; Ribonucleotide Reductases/metabolism; Thermodynamics; Tyrosine |
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