Fig. 1.

Obtaining the energy score of a model with an optimized orientation. The input orientation Ω_in is the Euler angle used to project the model, X. I_c is the centered target image, a constant parameter (step 1). A series of orientation angles Ω_k are generated around Ω_in (step 2, see SI Materials and Methods). The model X is projected along these proposed orientations to generate the corresponding projection images I_k (step 3). Negative cross-correlation scores are calculated between these projection images and the image I_c (step 4). The lowest value is assigned to the EM energy E_EM between the model X and the input image I_c (step 5). Its corresponding orientation is considered the optimized orientation Ω_opt that projects X to fit I_c. The outputs are the optimized projection orientation Ω_opt and the total energy E_total, which is the sum of a molecular energy, E_mol and the EM energy E_EM, weighted by an adjustable weight parameter w (step 6). E_mol is a function of the current model X to ensure its proper stereochemistry.