A possible glycine radical in anaerobic ribonucleotide reductase from Escherichia coli: nucleotide sequence of the cloned nrdD gene
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Summary
This summary is machine-generated.Researchers cloned the gene for an oxygen-sensitive ribonucleoside-triphosphate reductase in Escherichia coli, crucial for DNA replication during anaerobic growth. This enzyme shows similarity to a coliphage T4 gene, suggesting viral coding.
Area Of Science
- Molecular Biology
- Enzymology
- Genetics
Background
- Escherichia coli requires an oxygen-sensitive ribonucleoside-triphosphate reductase for DNA replication during anaerobic growth.
- This anaerobic enzyme is distinct from the aerobic ribonucleoside diphosphate-reductase (EC 1.17.4.1).
Purpose Of The Study
- To clone and characterize the gene encoding the anaerobic ribonucleoside-triphosphate reductase in E. coli.
- To investigate the functional and evolutionary relationship of this enzyme.
Main Methods
- Gene cloning of the anaerobic reductase.
- DNA sequencing to determine the coding region and identify regulatory elements (Fnr binding site).
- Amino acid sequence analysis and comparison with known proteins.
Main Results
- The cloned gene contains a 2136-nucleotide coding region (712 amino acids) with an Fnr binding site upstream.
- The deduced amino acid sequence exhibits 72% identity to coliphage T4's sunY gene, suggesting viral origin.
- A conserved pentapeptide motif (RVCGY) suggests Gly-681 as the site of the organic free radical, similar to pyruvate formate-lyase.
Conclusions
- The gene for the anaerobic ribonucleoside-triphosphate reductase has been identified and characterized in E. coli.
- The coliphage T4 sunY gene likely encodes a viral anaerobic reductase, indicating functional conservation.
- The study proposes a radical site on Gly-681, providing insights into the enzyme's activation mechanism.