Molecular Dynamics Simulations of the STAS Domains of Rat Prestin and Human Pendrin Reveal Conformational Motions in Conserved Flexible Regions

Abstract

Background: Molecular dynamics (MD) simulations provide valuable information on the conformational changes that accompany time-dependent motions in proteins. The reported crystal structure of rat prestin (PDB 3LLO) is remarkable for an α1-α2 inter-helical angle that differs substantially from those observed in bacterial STAS domains of SulP anion transporters and anti-sigma factor antagonists. However, NMR data on the rat prestin STAS domain in solution suggests dynamic features at or near the α1-α2 helical region (Pasqualetto et al JMB, 2010). We therefore performed a 100 ns 300K MD simulation study comparing the STAS domains of rat prestin and (modeled) human pendrin, to explore possible conformational flexibility in the region of the α1 and α2 helices. Methods: The conformation of the loop missing in the crystal structure of rat prestin STAS (11 amino acids between helix α1 and strand β3) was built using Modeller. MD simulations were performed with GROMACSv4.6 using GROMOS96 53a6 all-atom force field. Results: A subset of secondary structured elements of the STAS domains exhibits significant conformational changes during the simulation time course. The conformationally perturbed segments include the majority of loop regions, as well as the α1 and α2 helices. A significant decrease in the α1-α2 inter-helical angle observed across the simulation trajectory leads to closer helical packing at their C-termini. The end-simulation conformations of the prestin and pendrin STAS domains, including their decreased α1-α2 inter-helical angles, resemble more closely the packing of corresponding helices in the STAS structures of bacterial SulP transporters Rv1739c and ychM, as well as those of the anti-sigma factor antagonists. Several structural segments of the modeled human pendrin STAS domain exhibit larger atomic motions and greater conformational deviations than the corresponding regions of rat prestin, predicting that the human pendrin STAS domain in solution structure may be more dynamic than rat prestin STAS. Regions of prestin and pendrin identified by RMS fluctuation data as exhibiting larger atomic fluctuations corresponded to nominal GDP-binding regions of the aligned Rv1739c STAS domain of M. tuberculosis. Conclusions: MD simulations of mammalian STAS domains reveal substantial predicted conformational heterogeneity. These predicted conformational dynamics serve to supplement the reported crystal structure of the rat prestin STAS domain, and extend our understanding of the roles of STAS domains in SLC26 anion transporter function.