Computational and experimental mapping of the allosteric network of two manganese ABC transporters.

Transition metals (e.g., Fe, Zn, Mn) are essential enzymatic cofactors in all organisms. Their environmental scarcity led to the evolution of high-affinity uptake systems. Our research focuses on two bacterial manganese ABC importers, Streptococcus pneumoniae PsaBC and Bacillus anthracis MntBC, both critical for virulence. Both importers share a similar homodimeric structure, where each protomer comprises a transmembrane domain (TMD) linked to a cytoplasmic nucleotide-binding domain (NBD). Due to their size and slow turnover rates, the utility of conventional molecular simulation approaches to reveal functional dynamics is limited. Thus, we employed a novel, computationally efficient method integrating Gaussian Network Models (GNM) with information theory Transfer Entropy (TE) calculations. Our calculations are in remarkable agreement with previous functional studies. Furthermore, based on the calculations, we generated 10 point-mutations and experimentally tested their effects, finding excellent concordance between computational predictions and experimental results. We identified "allosteric hotspots" in both transporters, in the transmembrane translocation pathway, at the coupling helices linking the TMDs and NBDs, and in the ATP binding sites. In both PsaBC and MntBC, we observed bi-directional information flow between the two TMDs, with minimal allosteric transmission to the NBDs. Conversely, the NBDs exhibited almost no NBD-NBD allosteric crosstalk but showed pronounced information flow from the NBD of one protomer towards the TMD of the other protomer. This unique allosteric "footprint" distinguishes ABC importers of transition metals from other members of the ABC transporter superfamily establishing them as a distinct functional class. This study offers the first comprehensive insight into the conformational dynamics of these vital virulence determinants, providing potential avenues for developing urgently needed novel antibacterial agents.