Utilization of AlphaFold2 to Predict MFS Protein Conformations after Selective Mutation

Abstract

The major facilitator superfamily (MFS) is the largest secondary transporter family and is responsible for transporting a broad range of substrates across the biomembrane. These proteins are involved in a series of conformational changes during substrate transport. To decipher the transport mechanism, it is necessary to obtain structures of these different conformations. At present, great progress has been made in predicting protein structure based on coevolutionary information. In this study, AlphaFold2 was used to predict different conformational structures for 69 MFS transporters of E. coli after the selective mutation of residues at the interface between the N- and C-terminal domains. The predicted structures for these mutants had small RMSD values when compared to structures obtained using X-ray crystallography, which indicates that AlphaFold2 predicts the structure of MSF transporters with high accuracy. In addition, different conformations of other transporter family proteins have been successfully predicted based on mutation methods. This study provides a structural basis to study the transporting mechanism of the MFS transporters and a method to probe dynamic conformation changes of transporter family proteins when performing their function.

Accelerated Molecular Dynamics and AlphaFold Uncover a Missing Conformational State of Transporter Protein OxlT

“…Although previous studies suggested that AF2 is unable to predict point mutations that cause misfolding or destabilization of the native structure, , the situation is different for the protein conformational change because AF2 can learn alternative conformations of homologous proteins in PDB. A previous study also showed a possibility that two or more mutations allow AF2 to explore an alternative conformation of transporter proteins . In combination with the accelerated MD simulation, we here clarified that even a single mutation in AF2 can shift the predicted conformation.…”

“…A previous study also showed a possibility that two or more mutations allow AF2 to explore an alternative conformation of transporter proteins. 42 In combination with the accelerated MD simulation, we here clarified that even a single mutation in AF2 can shift the predicted conformation. Furthermore, we showed that AF2 prediction is affected by the input MSA, and residuewise coevolution score derived from the MSA can pin down the state-shifting mutations.…”

Nramp: Deprive and conquer?

“…Mining sites of interaction/flexibility ( Zhang et al, 2020 ) that can be mutated to produce, or induce AF2 modeling of, alternate conformers ( Bozzi et al, 2016a ; Bozzi et al, 2016b ; Del Alamo et al, 2022 ; Stein and Mchaourab, 2022 ; Xiao et al, 2022 ) should be useful to represent the spectrum of conformational diversity in the Slc11 family, and comparative analyses across phylogroups will aid probing the dynamics of Slc11 carrier cycle and its plasticity throughout evolution.…”

Accelerated Molecular Dynamics and AlphaFold Uncover a Missing Conformational State of Transporter Protein OxlT

“…A previous study also showed a possibility that two or more mutations allow AF2 to explore an alternative conformation of transporter proteins. 38 In combination with the accelerated MD simulation, we here clarified that even a single mutation can shift AF2-predicted conformation. Furthermore, we showed that AF2 prediction is affected by the input MSA, and residue-wise coevolution score derived from the MSA can pin down the state- shifting mutations.…”