Ribonucleotide reductase in the archaeon Pyrococcus furiosus: a critical enzyme in the evolution of DNA genomes?

Abstract

Ribonucleotide reductase (RNR), the enzyme responsible for deoxyribonucleotide synthesis, has been isolated from Pyrococcus furiosus, a deeply branching hyperthermophilic, strictly anaerobic archaeon. Its gene has been cloned, sequenced, and shown to harbor two insertions encoding inteins. The purified enzyme absolutely requires adenosylcobalamin for activity, a trait that defines it as a member of class II (adenosyl-cobalamin-dependent) prokaryotic RNRs. On the other hand, the archaeal RNR has significant amino acid sequence homology with class I (aerobic non-heme iron-dependent) and class III (anaerobic iron-sulfur-dependent) RNRs present in eukaryotes and bacteria, respectively. It is proposed that this enzyme may be the closest possible relative of the original RNR, which allowed the key "RNA world" to "DNA world" transition, and that the different classes of present-day RNRs are the products of divergent evolution.

Figure 1

(a) Comparison of the N-terminal sequences of P. furiosus RNR, the large subunit of class I (mouse) RNR, and the large subunit of class III (anaerobic E. coli) RNR. (b) Alignment of the predicted amino acid sequences of P. furiosus (upper sequence) and mouse (large subunit) RNRs using the blastx program. The two inteins of the archaeal RNR are double-underlined. The conserved LAGLI-DADG motif characteristic of protein spacer regions is indicated with the symbol ∗ (7). Critical amino acids at the intein–extein junctions are indicated with the symbol #. The entire RNR sequence was determined by dideoxy sequencing of a 9-kb plasmid subclone from a P. furiosus genomic library (R.W., D. M. Dunn, J. L. Cherry, M. D. Stump, P. Cartwright, et al. unpublished work).

Figure 2

The essential cysteines in the large subunit of class I (mouse) RNR and in class II (L. leichmannii) RNR are present in P. furiosus RNR. C₁ and C₃ designate the cysteines that directly reduce the ribonucleotide. C₂ is the cysteinyl radical (13).