Structure-Based Strategy Reveals a Novel Ligand Binding Site in Staphylococcus aureus Catabolite Control Protein A:...

Staphylococcus aureus is a major human pathogen responsible for a broad spectrum of infections and is recognized by the World Health Organization as a high-priority antimicrobial-resistant organism. The global transcription factor (TF) catabolite control protein A (CcpA), a central regulator of metabolism and virulence, represents a promising drug target. However, efforts to design inhibitors are hindered by their closed crystallographic conformation and the absence of well-defined ligand-binding cavities. Here, we present a structure-guided alignment strategy to identify potential ligand-binding sites in CcpA from S. aureus (Sa-CcpA), using a curated set of cocrystallized TFs as structural templates. This methodology identified a putative ligand-binding site in Sa-CcpA that was not predicted by conventional cavity-detection methods. Phylogenetic analysis showed that residues within this predicted cavity are conserved in a subset of particular pathogenic species in the order Bacillales, suggesting an evolutionarily constrained and functionally relevant region. To assess the stability of this newly identified site, multiple molecular dynamics simulations were performed, followed by docking analyses of two known Sa-CcpA inhibitors. Together, our results suggest that this structure-informed alignment strategy is a promising approach to uncover previously unrecognized ligand-binding regions in TFs. Although demonstrated here using Sa-CcpA as a case study, this approach may support future efforts to rationalize inhibitor design targeting transcriptional regulators.