Accurate Protein Dynamic Conformational Ensembles: Combining AlphaFold, MD and Amide 15N(1H) NMR Relaxation.

Conformational heterogeneity is critical for protein function, but the validation of dynamic ensembles remains a challenge. In this study, we introduced an approach that integrates free MD simulations, using an AlphaFold-generated structure as the starting point, with experimental relaxation data to identify biologically relevant conformational ensembles. For the extracellular region of Streptococcus pneumoniae PsrSp, we found that only certain segments of the MD long trajectory aligned well with experimental data. The defined ensembles revealed two regions with increased flexibility that play important functional roles.