X-ray crystallographic studies of the archaeal holliday junction resolvase HJC from A. fulgidus and its complex with a 4-way DNA junction

Holliday junction-resolving enzymes (X-resolvases) mediate the reaction by recognizing DNA four-way junctions and introducing symmetrical nicks in a very precise manner. They are dimeric proteins that recognize DNA four-way junctions and resolve them with high accuracy by introducing symmetrical nicks on both sides of the branch point. X-resolvases exhibit a stretch of basic amino acids on their surface that are responsible for DNA binding. I have set out to study the structure of the archaeal resolvase Holliday junction resolvase from Archaeoglobus fulgidus, alone and in complex with 4-way junction, using X-ray crystallography. The crystal structure of Hjc alone was solved at 1.63 Å resolution, and it shows the same overall fold of other Hjc family members. The structure was not complete, due to the distortion of some parts. In order to obtain the complete structure, a surface mutant was created by site mutagenesis of two non-conserved residues. The surface mutant structure, solved at 1.56 Å, shows a complete protein, and the catalytic flexible loop can be observed distorted in two different functional conformations.I also report the crystal structure of Hjc in complex with the 4-way junction with 10bp long arms, at 3.15 Å resolution. This structure provides the first picture of a Holliday junction bound to the Holliday junction cutting enzyme in archaea. The structural model shows a novel form of protein- Holliday junction binding, the biological meaning of which is yet to be clarified.