Interactions of glutaredoxins, ribonucleotide reductase, and components of the DNA replication system of Escherichia coli.
A strain of Escherichia coli missing three members of the thioredoxin superfamily, thioredoxins 1 and 2 and glutaredoxin 1, is unable to grow, a phenotype presumed to be due to the inability of cells to reduce the essential enzyme ribonucleotide reductase. Two classes of mutations can restore growth to such a strain. First, we have isolated a collection of mutations in the gene for the protein glutaredoxin 3 that suppress the growth defect. Remarkably, all eight independent mutations alter the same amino acid, methionine-43, changing it to valine, isoleucine, or leucine. From the position of the amino acid changes and their effects, we propose that these alterations change the protein so that its properties are closer to those of glutaredoxin 1. The second means of suppressing the growth defects of the multiply mutant strain was by mutations in the DNA replication genes, dnaA and dnaN. These mutations substantially increase the expression of ribonucleotide reductase, most likely by altering the interaction of the regulatory protein DnaA with the ribonucleotide reductase promoter. Our results suggest that this increase in the concentration of ribonucleotide reductase in the cell allows more effective interaction with glutaredoxin 3, thus restoring an effective pool of deoxyribonucleotides. Our studies present direct evidence that ribonucleotide reductase is the only essential enzyme that requires the three reductive proteins missing in our strains. Our results also suggest an unexpected regulatory interaction between the DnaA and DnaN proteins.
Ortenberg
Ron
R
Department of Microbiology and Molecular Genetics, Harvard Medical School, 200 Longwood Avenue, Boston, MA 02115, USA.
Gon
Stéphanie
S
Porat
Amir
A
Beckwith
Jon
J
D000595:Amino Acid Sequence; D004261:DNA Replication; D004926:Escherichia coli / Q000235:genetics / Q000254:growth & development / Q000378:metabolism*; D054477:Glutaredoxins; D008969:Molecular Sequence Data; D009154:Mutation; D010088:Oxidoreductases / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding; D012264:Ribonucleotide Reductases / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid
D054477:Glutaredoxins; D010088:Oxidoreductases; D012264:Ribonucleotide Reductases