Molecular Insights into the Allosteric Regulation of Pyruvate Kinase in Streptococcus pneumoniae

Streptococcus pneumoniae is a clinically important fermentative pathogen that relies heavily on glycolysis for energy production. Its rate-limiting enzyme, pyruvate kinase (SpPYK), is allosterically activated by the effector fructose-1,6-bisphosphate that binds ∼40 Å away from the substrate-binding catalytic site; however, the dynamic mechanism driving this long-range communication remains elusive. In this study, we employ molecular dynamics (MD) simulations and network analysis to map the allosteric activation pathway of SpPYK. Our results indicate that effector binding reshapes the dynamical landscape of the protein, increasing the coherence of collective motions coupled to active-site activation and rewiring long-range correlations. Our computational findings were validated by mutagenesis assays, which confirmed the role of key leucine residues within the activation pathway for the catalytic activity of the enzyme. Our findings refine the “Rock and Lock” model of allostery and offer a structural blueprint for leveraging allosteric regulation to develop species-selective antimicrobials.