Homology modelling of Bim1

A homology model of Bim1 was created using the one to one threading option in Phyre2⁵⁴, based on the tetragonal crystal structure of LaX325³², PDB code 6IBJ. Bim1 and LaX325 share 36% sequence identity. Local alignment and secondary structure scoring (with a weight of 0.1) were used for producing a model with 100% Phyre2 confidence (this just indicates the confidence that the two sequences aligned truly represent two homologous proteins). Phyre2 aligns Hidden Markov Models (HMMs) of query and target. Based on this alignment, Phyre2 builds a crude backbone model based on the template, then fills missing loops using a fragment database from known structures. Finally, side chains were built in their most probable rotamer while avoiding clashes. In order to improve the geometry of the model, two Ramachandran outliers were corrected in COOT⁵⁵, and 20 cycles of structure idealization carried out in REFMAC5⁵⁶ after adding coordinates for Cu²⁺ based on the template structure. The final model had good geometry as evaluated using the Molprobity server⁵⁷. The Cα RMSD between the Bim1 model and the template was 0.679 Å.