Assessing the utility of CASP14 models for molecular replacement

Abstract

The assessment of CASP models for utility in molecular replacement is a measure of their use in a valuable real-world application. In CASP7, the metric for molecular replacement assessment involved full likelihood-based molecular replacement searches; however, this restricted the assessable targets to crystal structures with only one copy of the target in the asymmetric unit, and to those where the search found the correct pose. In CASP10, full molecular replacement searches were replaced by likelihood-based rigid-body refinement of models superimposed on the target using the LGA algorithm, with the metric being the refined likelihood (LLG) score. This enabled multi-copy targets and very poor models to be evaluated, but a significant further issue remained: the requirement of diffraction data for assessment. We introduce here the relative-expected-LLG (reLLG), which is independent of diffraction data. This reLLG is also independent of any crystal form, and can be calculated regardless of the source of the target, be it X-ray, NMR or cryo-EM. We calibrate the reLLG against the LLG for targets in CASP14, showing that it is a robust measure of both model and group ranking. Like the LLG, the reLLG shows that accurate coordinate error estimates add substantial value to predicted models. We find that refinement by CASP groups can often convert an inadequate initial model into a successful MR search model. Consistent with findings from others, we show that the AlphaFold2 models are sufficiently good, and reliably so, to surpass other current model generation strategies for attempting molecular replacement phasing.

Using AlphaFold to predict the impact of single mutations on protein stability and function

“…These problems include the prediction of various protein interactions, such as protein-protein, proteinligand and protein-DNA/RNA, and the prediction of the impact of mutations on protein stability. AlphaFold proved to be useful for experimental determination of protein structures with molecular replacement phasing [4,5] and already facilitated elucidation of SARS-Cov2 protein structures [6,7]. Furthermore, AlphaFold in collaboration with EMBL-EBI launched a global initiative on constructing the structure models for the whole protein sequence space [8].…”

Structural and Functional Studies on Evolutionary Repurposing of H-box/NC-proteins : From Host Factor to Virus Protein

“…AlphaFold2. The availability and accuracy of predicted structures has increased since the release of AlphaFold2, making molecular replacement the first choice to obtain an initial set of phases (31). Especially since Al-phaFold2 can predict protein models solely from the protein sequence and can then be used for almost any protein independent of prior known structural data.…”

“…Computational methods can be extremely useful for experimentalists e.g. as mentioned above where AF2 today has become the go-to approach to help solve the "phase problem" in protein crystallography by providing accurate search models for molecular replacement (31). Additionally, AF2 can be used to build a hypothesis around protein functions and interactions (38).…”