Prediction of protein assemblies by structure sampling followed by interface‐focused scoring

Proteins often function as part of permanent or transient multimeric complexes, and understanding function of these assemblies requires knowledge of their three-dimensional structures. While the ability of AlphaFold to predict structures of individual proteins with unprecedented accuracy has revolutionized structural biology, modeling structures of protein assemblies remains challenging. To address this challenge, we developed a protocol for predicting structures of protein complexes involving model sampling followed by scoring focused on the subunit-subunit interaction interface. In this protocol, we diversified AlphaFold models by varying construction and pairing of multiple sequence alignments as well as increasing the number of recycles. In cases when AlphaFold failed to assemble a full protein complex or produced unreliable results, additional diverse models were constructed by docking of monomers or subcomplexes. All the models were then scored using a newly developed method, VoroIF-jury, which relies only on structural information. Notably, VoroIF-jury is independent of AlphaFold self-assessment scores and therefore can be used to rank models originating from different structure prediction methods. We tested our protocol in CASP15 and obtained top results, significantly outperforming the standard AlphaFold-Multimer pipeline. Analysis of our results showed that the accuracy of our assembly models was capped mainly by structure sampling rather than model scoring. This observation suggests that better sampling, especially for the antibody-antigen complexes, may lead to further improvement. Our protocol is expected to be useful for modeling and/or scoring protein assemblies.