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

ABSTRACT

Transporter proteins change their conformations to carry their substrate across the cell membrane. The conformational dynamics is vital to understanding the transport function. We have studied the oxalate transporter (OxlT), an oxalateformate antiporter from Oxalobacter formigenes, significant in avoiding kidney stone formation. The atomic structure of OxlT has been recently solved in the outward-open and occluded states. However, the inward-open conformation is still missing, hindering a complete understanding of the transporter. Here, we performed a Gaussian accelerated molecular dynamics simulation to sample the extensive conformational space of OxlT and successfully predicted the inward-open conformation where cytoplasmic substrate formate binding was preferred over oxalate binding. We also identified critical interactions for the inward-open conformation. The results were complemented by an AlphaFold2 structure prediction. Although AlphaFold2 solely predicted OxlT in the outward-open conformation, mutation of the identified critical residues made it partly predict the inward-open conformation, identifying possible state-shifting mutations.