Abstract
A major goal in structural biology is to understand protein assemblies in their biologically relevant states. Here, we investigate whether AlphaFold2 structure predictions match native protein conformations. We chemically cross-linked proteins in situ within intact Tetrahymena thermophila cilia and native ciliary extracts and identified 1,225 intramolecular cross-links within the 100 best-sampled proteins to provide a benchmark of distance restraints obeyed by proteins in their native assemblies. The corresponding AlphaFold2 structure predictions were highly concordant, positioning 86.2% of cross-linked residues within Cα-to-Cα distances of 30 Å, consistent with the known cross-linker length. 43% of the proteins showed no violations. Most inconsistencies occurred in low-confidence regions or between domains of the structure prediction. For basal body protein BBC118, cross-links combined with the predicted structure revealed domain packing satisfying both data. Overall, AlphaFold2 predicted biological structures with low predicted aligned error corresponded to more correct native structures. However, we observe cases where rigid body domains are oriented incorrectly, suggesting that combining structure prediction with experimental information will better reveal biologically relevant conformations.
Competing Interest Statement
The authors have declared no competing interest.
