We can rule out overfitting for three reasons: (1) Substantial portions of the model were not built from scratch, but they were rigid-body fitted domains from our 2.4 Å x-ray structure of soluble PLY. These map regions cannot be overfitted. (2) The major new part of the membrane-inserted form of the PLY protomer is a β-barrel with 84 long β-hairpins of rigid, pre-defined geometry, so the risk of overfitting in this region is minimal. (3) The comparison of the phase-randomized vs masked FSC (Figure 1—figure supplement 2B) indicates that there is no overfitted noise.