LAUSR

RecJ is ubiquitous in bacteria and Archaea, and play an important role in DNA replication and repair. Currently, our understanding on biochemical function of archaeal RecJ is incomplete due to limited reports. The genome of the hyperthermophilic and radioresistant euryarchaeon Thermococcus gammatolerans encodes one putative RecJ protein (Tga-RecJ). Herein, we report biochemical characteristics and catalytic mechanism of Tga-RecJ. Tga-RecJ can degrade ssDNA in the 5'-3' direction at high temperature as observed in Thermococcus kodakarensis RecJ and Pyrococcus furiosus RecJ, the two closest homologs of the enzyme. In contrasted to P. furiosus RecJ, Tga-RecJ lacks 3'-5' ssRNA exonuclease activity. Furthermore, maximum activity of Tga-RecJ is observed at 50 °C ~ 70 °C and pH 7.0-9.0 with Mn2+, and the enzyme is the most thermostable among the reported RecJ proteins. Additionally, the rates for hydrolyzing ssDNA by Tga-RecJ were estimated by kinetic analyses at 50 °C ~ 70 °C, thus revealing its activation energy (10.1 ± 0.6 kcal/mol), which is the first report on energy barrier for ssDNA degradation by RecJ. Mutational studies showed that the mutations of residues D36, D83, D105, H106, H107 and D166 in Tga-RecJ to alanine almost completely abolish its activity, thereby suggesting that these residues are essential for catalysis.