INTERACTIONS OF 2'-MODIFIED AZIDOANALOGS AND HALOANALOGS OF DEOXYCYTIDINE 5'-TRIPHOSPHATE WITH THE ANAEROBIC RIBONUCLEOTIDE REDUCTASE OF ESCHERICHIA-COLI

The anaerobic Escherichia coli ribonucleotide reductase (class III red uctase) responsible for the synthesis of the deoxyribonucleotides requ ired for anaerobic DNA replication contains an oxygen-sensitive glycyl radical (Gly-681) suggesting involvement of radical chemistry in cata lysis. The amino acid sequence of this enzyme completely differs from that of earlier described aerobic class I (prototype, aerobic E. coli) and class II (proto type, Lactobacillus leichmanii) reductases that u se radical chemistry but employ other means for radical generation. He re, we study the interaction between the anaerobic E. coli reductase w ith the 5'-triphosphates of 2'-chloro-2'-deoxycytidine, 2'-fluoro-2'-d eoxycytidine, and 2'-azido-2' deoxycytidine (N3CTP), which are mechani sm-based inhibitors of class I and II reductases and, on interaction w ith these enzymes, decompose to base, inorganic di(tri)phosphate and 2 '-methylene-3(2H)-furanone. Also, with the anaerobic E. coli reductase , the 2'-substituted nucleotides act as mechanism-based inhibitors and decompose. N3CTP scavenges the glycyl radical of the enzyme similar t o the interaction of N3CDP with the tyrosyl radical of class I enzymes . However we found no evidence for a new transient radical species as is the case with class I enzymes. Our results suggest that the chemist ry at the nucleotide level for the reduction of ribose by class III en zymes is similar to the chemistry employed by class I and II enzymes.