Hernández-Escamilla R, Espinosa-Lara JM, Quintana-Hau JD, Uribe-Luna S,

Language: English
References: 27
Page: 58-64
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ABSTRACT

The aim of this work was to search if the rat DNA polymerase beta can substitute the capability of DNA polymerase I to repair damage caused by the UV light in Escherichia coli. The oriC origin of replication from pβ5 was replaced by the rep origin from pSC101 and named pβ6. The presence of pol beta in the new construct was verified by PCR. E. coli polA-1 (WP6) was transformed with pβ6. A protein with size similar to DNA Pol beta (40 kDa) was shown in the cell free extracts carrying pβ5. In WP6/pβ6 cell free extracts a slightly smaller protein was observed instead of the 40 kDa. DNA Pol beta was revealed by western analysis, with polyclonal antibodies, in strains with pβ5. Yet, it was not detected in the western from WP6/pβ6. A moderate change in UV resistance was observed in strains carrying pβ5. However, in polA1 carrying pΒ6 (WP6/pβ6), irradiated with 60-90 J/m² of UV light, the viability was increased by more than four orders of magnitude, when compared with the polA1 (WP6) strain, reaching approximately the same UV resistance as the strains with DNA polymerase I. The results suggests that probably Pol beta is rapidly degraded in the cell free extracts from WP6/pβ6 and, it repairs the lethal effect of the UV light in E. coli.

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