An atomic model was built and refined against the GagΔMAT8I density map. The crystal structure (PDB 4XFX)56 and cryoEM structure (PDB 5L93)11 of HIV-1 CA were used as an initial reference. At 4.2 Å resolution, positive and bulky side chains are clearly visible enabling reasonable positioning of the residues in the atomic models. Rotamers were generally not refined, unless there was good evidence for a different rotamer in our density map. For the SP1 region, a composite model was made by joining the SP1 residues from PDB 1U579 to the 5L93 model (lacking the last 6 SP1 residues) to serve as initial reference for the model refinement.

The model refinement procedure involved iterative manual refinement in Coot57 followed by three rounds of Phenix cryoEM real-space refinement58. After each round of real-space refinement, Ramachandran and rotamer outliers were manually refined in Coot, and another round of real-space refinement was performed until good model statistics were achieved. In order to ensure reasonable modelling in the SP1 region, the density map of GagΔMAT8I was sharpened with a B factor of 120 in combination with a 1% FDR confidence map34 and a locally sharpened B-factor map59 for the manual refinement in Coot.

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