Computational protocol: PARP-2 domain requirements for DNA damage-dependent activation and localization to sites of DNA damage

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Protocol publication

[…] Crystals were grown by sitting drop vapor diffusion at room temperature in 0.6–0.7 M sodium citrate, 0.1 M sodium citrate buffer pH 5.6 and 7–10 mM DTT, and the protein complex at 15 mg/ml. Crystals were cryoprotected in reservoir solution supplemented with 25% glycerol (or ethylene glycol) and then flash-cooled in liquid nitrogen. Diffraction data were collected at beamline X29 at the National Synchrotron Light Source (NSLS, Brookhaven National Laboratory). The data were processed using Xia2 and XDS (,). The initial set of phases came from rigid body refinement of mouse importin α1 ΔIBB (PDB code 1Y2A, with the bound NLS removed from the structure). The human PARP-2 NTR NLS was built manually in COOT (). Iterative rounds of refinement were performed using PHENIX () and REFMAC in the CCP4 Suite of programs (–). […]

Pipeline specifications

Software tools xia2, XDS, Coot, PHENIX, CCP4
Applications Small-angle scattering, Protein structure analysis
Organisms Homo sapiens
Chemicals Poly Adenosine Diphosphate Ribose