The antimutagenic effect of a truncated epsilon subunit of DNA polymerase 3 in Escherichia coli cells irradiated with UV light
1995
Kanabus, M. | Nowicka, A. | Sledziewska-Gojska, E. | Jonczyk, P. | Ciesla, Z. (Polish Academy of Sciences, Warsaw (Poland). Inst. of Biochemistry and Biophysics)
It has previously been suggested that inhibition of the proofreading 3'-5' exonuclease activity of DNA polymerase may play an important role in generation of UV-induced mutations in Escherichia coli. Previous work showing that overproduction of epsilon, the proofreading subunit of DNA polymerase 3, counteracts the SOS mutagenic response of E. coli seemed to be consistent with this hypothesis. To explore further the nature of the antimutagenic effect of epsilon plasmid pMK17 was constructed, which encodes only two of the three highly conserved segments of epsilon -Exo1 and Exo2; the third segment, Exo3, which is essential for 3'-5' exonuclease activity, is deleted. At 40 degrees C, overproduction of the truncated epsilon subunit significantly delays production of M13 phage, suggesting that the protein retains its capacity to bind to DNA. On the other hand, the presence of pMK17 in a trpE65 strain growing at 40 degrees C causes a 10-fold decrease in the frequency of UV-induced Trp(+) mutations. This antimutagenic effect of the truncated epsilon is effectively relieved by excess UmuD,C proteins. The presence of plasmid pIP21, which contains the dnaQ49 allele encoding an epsilon subunit that is defective in proofreading activity, almost completely prevents generation of UV-induced mutations in the trpE65 strain. It is proposed that the DNA binding ability of free epsilon, rather than its 3'-5' exonuclease activity, affects processing of premutagenic UV-induced lesions, possibly by interfering with the interaction between the UmuC -UmuD'-RecA complex and Pol 3 holoenzyme. This interaction is probably a necessary condition for translesion synthesis.
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