Efficient DNA damage repair by a novel photolyase from Acinetobacter sp. Ver3, a UV-resistant extremophile.

SIMON, J.; ALBARRACÍN V. H.; FARIAS M.E.; GÄRTNER W.

Leipzig

Congreso; 5RD FEMS CONGRESS OF EUROPEAN MICROBIOLOGISTS; 2013

FEMS

The extremophilic Acinetobacter sp. Ver3 was previously described as an exceptionally UV-resistant strain, isolated from Lake Verde (4,400 m asl), a hypersaline lake in Argentinean Puna. Evidences point out to this interesting UV-resistance phenotype caused by efficient DNA photorepairing ability, coupled to highly efficient antioxidant enzyme activities in response to UV-B stress. We herein present the functional characterization of a CPD-Class I photolyase (Ver3Phr) from Acinetobacter sp. Ver3 to test its involvement on the previously observed photoreactivation ability. UV-vis and fluorescence spectroscopy of the overexpressed and purified protein identified a FAD chromophore and a MTHF (methyltetrahydrofolate) antenna. Functional in vitro tests demonstrated repair capability of Ver3Phr on a UV-damaged poly-dT-oligonucleotide. Also, functional complementation analysis of the Phr gene of Acinetobacter sp. Ver3 using photoreactivation deficient E. coli KY1225 cells was performed. Transformants carrying the pQE60-Ver3Phr or the controls without plasmid were UV-B irradiated. When plated immediately after treatment, it was not observed any cell survival in both cases. E. coli strains were then exposed to light (PR) or dark repair (DR). Non-transformant KY1225 cells showed not recovery at all after both treatments. When photoreactivated, cells carrying Ver3Phr reached almost the same CFU numbers than non-UV treated cells while under dark conditions a much minor recovery in cell growth was observed: i.e. six orders of magnitude lower than under PR. These results indicated that the protein encoded by the Phr gene of Acinetobacter sp. Ver3 has a photoreactivation activity while its contribution to dark repair is practically insignificant.