| Budget Amount *help |
¥3,500,000 (Direct Cost: ¥3,500,000)
Fiscal Year 2001: ¥1,400,000 (Direct Cost: ¥1,400,000)
Fiscal Year 2000: ¥2,100,000 (Direct Cost: ¥2,100,000)
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| Research Abstract |
RecJ is a 5' to 3' exonuclease specific for single-stranded DNA, and involved in DNA repair and recombination systems. Recently, RecJ has been highlighted as a key enzyme for the replication-recombination relationship. RecJ homologues have five characteristic motifs, which form a large family of the predicted phosphoesterases, named by DHH family.
We performed the X-ray crystallography of RecJ from Thermus thermohilus HB8 (ttRecJ).
As ttRecJ was expressed as an inclusion body, it was purified through denaturation and refolding. The region of the catalytic core domain (cd-ttRecJ) was identified by limited proteolysis. Cd-ttRecJ, expressed as soluble form, was highly purified and crystallized.
The structure of RecJ was solved by single-wavelength anomalous dispersion, using Se-Met cd-ttRecJ. RecJ has a novel fold, in which two domains are interconnected with a long helix to form a central groove. This groove is composed of conserved residues and positively charged, which may be involved in DNA binding. The metal ion is coordinated by the motifs characteristic in DHH family. This metal ion was identified as Mn^<2+> by an anomalous scattering experiment.
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