Adaptive repair of cross-links in DNA of Micrococcus radiodurans

Cross-links in the DNA of Micrococcus radiodurans induced by mitomycin C were repaired during post-incubation. This repair process was inhibited in cells post-incubated in the presence of chloramphenicol. However, the removal of cross-links in DNA was almost normal, even in the presence of chloramphenicol, if the cells were pretreated with lower concentrations of mitomycin C.     

DNA-membrane complex restoration in Micrococcus radiodurans after X-irradiation: relation to repair, DNA synthesis and DNA degradation

The DNA-membrane complex in Micrococcus radiodurans was shown to be essentially constituted of proteins, lipids and DNA. The complex was dissociated immediately after X-irradiation of cells and restored during post-incubation in complete medium. In X-irradiated protoplasts some DNA remained associated with the complex. Restoration of the complex during post-incubation was only seen in a medium favouring DNA polymerase and ligase activities. Under this condition no DNA synthesis occurred, suggesting that complex restoration may involve ligase activity. The complex restoration in the wild type and the X-ray sensitive mutant UV17 of M. radiodurans was strictly dependent on the X-ray dose. It was correlated with survival and DNA degradation but always preceded the onset of DNA synthesis after X-irradiation. At the same dose the complex restoration was about 2 fold lower in mutant than in wild type cells indicating that the restoration of the complex is related to repair capacity. The results are consistent with the idea that the complex protects X-irradiated DNA of M. radiodurans from further breakdown and, subsequently, permits DNA synthesis and repair to occur.     

Separation of DNA-dependent DNA polymerase activities in Micrococcus radiodurans.

DNA polymerase activities in Micrococcus radiodurans were separated into two fractions after purification more than 2000 fold. They differ in pH optimum and residual activities in the absence of a full deoxyribonucleoside triphosphates complement. NAD partly inhibited one of the activities. Both activities were eluted as a single peak on gel filtration and sedimented at the same rate on glycerol gradient centrifugation. Molecular weight 140000 was calculated from Stokes radius and sedimentation constant. Deoxyribonuclease activity was detected on one of the polymerase activities which preferentially degraded double-stranded DNA. Priming activity of nicked DNA was reduced by gamma-irradiation. These results have been related to the possible rolls in repair synthesis in vivo or DNA synthesis in permeable cells of M. radiodurans.     

In situ enzymology of DNA replication and ultraviolet-induced DNA repair synthesis in permeable human cells

Using permeable diploid human fibroblasts, we have studied the deoxyribonucleoside triphosphate concentration dependences of ultraviolet- (UV-) induced DNA repair synthesis and semiconservative DNA replication. In both cell types (AG1518 and IMR-90) examined, the apparent Km values for dCTP, dGTP, and dTTP for DNA replication were between 1.2 and 2.9 microM. For UV-induced DNA repair synthesis, the apparent Km values were substantially lower, ranging from 0.11 to 0.44 microM for AG1518 cells and from 0.06 to 0.24 microM for IMR-90 cells. Control experiments established that these values were not significantly influenced by nucleotide degradation during the permeable cell incubations or by the presence of residual endogenous nucleotides within the permeable cells. Recent data implicate DNA polymerase delta in UV-induced repair synthesis and suggest that DNA polymerases alpha and delta are both involved in semiconservative replication. We measured Km values for dGTP and dTTP for polymerases alpha and delta, for comparison with the values for replication and repair synthesis. Km values for polymerase alpha were 2.0 microM for dGTP and 5.0 microM for dTTP. For polymerase delta, the Km values were 2.0 microM for dGTP and 3.5 microM for dTTP. The deoxyribonucleotide Km values for DNA polymerase delta are much greater than the Km values for UV-induced repair synthesis, suggesting that when polymerase delta functions in DNA repair, its characteristics are altered substantially either by association with accessory proteins or by direct posttranslational modification. In contrast, the deoxyribonucleotide binding characteristics of the DNA replication machinery differ little from those of the isolated DNA polymerases.(ABSTRACT TRUNCATED AT 250 WORDS)     

Lysis of Micrococcus radiodurans

Two sulfur containing peptides have been isolated from a cationic peptide-rich fraction of dried Lentinus edodes by Chromatographic technic. They have been identified as γ-glutamylcystine and N,N-bis-³-glutamylcystinylglycine based on chemical, Chromatographic and spectroscopic properties.     

Genetic control of the processes of postradiation repair of a compact chromosome in Micrococcus radiodurans cells

X-irradiation of Micrococcus radiodurans cells with sublethal doses caused disturbances in the structure of a membrane-bound compact chromosome. Recovery of the compact chromosome occurred during the postirradiation incubation of the wild type cells and cells of the UVS-17 mutant deficient in DNA-polymerase. This process was blocked in cells of rec-30 mutant with the impaired system of genetic recombination: this is indicative of an important role played by rec-30 gene product in the postirradiation recovery of the compact chromosome in M. radiodurans cells.     

Radioresistance mechanisms of Micrococcus radiodurans

The modern conceptions on the molecular mechanisms of Micrococcus radiodurans viability under the action of ionized radiation have been considered. Factors providing a high level of the bacterium radioresistance-the peculiarities of the cell wall structure, membranes, DNA, the redundancy of genetic information, the multiplicity of sites of DNA attachment to the membrane, a high level of antioxidant and antiradical systems-have been analysed. It has been shown that the efficiency of accurate, error-free, well balanced DNA repair system in connection with M. radiodurans properties mentioned provides a high radiation resistance of this microorganism.     

ADP effects on bleomycin-induced DNA repair synthesis and adenylate kinase activity in permeable mouse sarcoma cells

DNA repair in bleomycin-pretreated, permeable mouse sarcoma (SR-C3H/He) cells requires ATP for at least two steps, the repair DNA synthesis step and the repair patch ligation step. ADP can apparently replace ATP in both steps. Maximal, 1.5-2 fold stimulation of repair DNA synthesis was observed with 5-10 mM ADP as well as 2.5-5 mM ATP. Repair patch ligation in the presence of 2.5 mM ADP occurred at almost the same high efficiency as it did in the presence of ATP. The ADP effect on DNA repair patch ligation was attributed to ATP formed from ADP by adenylate kinase in permeable cells, however the ADP effect on repair DNA synthesis could not be attributed solely to the formation of ATP in the same manner.     

Antibiotic Sensitivity of Micrococcus radiodurans

A wild-type strain of Micrococcus radiodurans and its nonpigmented mutant W(1) were tested for sensitivity to 10 antibiotics selected from the standpoint of their mechanism of action. Representatives of groups of antibiotics inhibiting deoxyribonucleic acid (DNA) synthesis, DNA-dependent ribonucleic acid synthesis, protein synthesis, and cell wall synthesis were selected. M. radiodurans and its mutant exhibited full susceptibility to all antibiotics tested (mitomycin C, actinomycin D, chloramphenicol, dihydrostreptomycin, erythromycin, neomycin, kanamycin, benzylpenicillin, bacitracin, and vancomycin), the degree of susceptibility being of the same order as that of a standard strain of Staphylococcus aureus 209 P, with the exception of dihydrostreptomycin.