Computational protocol: Insights into DNA substrate selection by APOBEC3G from structural, biochemical, and functional studies

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Protocol publication

[…] There were four Pot1-A3GCTD molecules in the asymmetric unit of the crystal. The structure was solved by molecular replacement using PHASER [] with the A3GCTD structure (PDBID 3IR2 []) and the Pot1 structure (PDBID 1QZH []) as search models. Clear electron density of Pot1-A3GCTD, including that for the Pot1 cognate DNA, was evident in the electron density map. Additional electron density was observed for the adenine at the 5’ side of the Pot1 cognate DNA sequence. Furthermore, weak electron density 5’ to the density was also observed but the quality was not sufficient for model building. Rigid body and iterative rounds of restrained refinement (including amplitude-based twin refinement) were carried out using Refmac5 [], followed by rebuilding the model to the 2Fo-Fc and the Fo-Fc maps using Coot []. Non-crystallographic symmetry restraints were applied in the refinement cycles. The final model has an Rwork/Rfree of 23.3%/28.9%. The refinement statistics are summarized in . The structure was analyzed and illustrated with Coot and PyMOL []. […]

Pipeline specifications

Software tools REFMAC5, Coot, PyMOL
Application Protein structure analysis
Organisms Homo sapiens
Chemicals Adenine, Cytidine, Deoxycytidine, Deoxyuridine, Nucleotides