Electrostatic-Mediated Affinity Tuning of Lysostaphin Accelerates Bacterial Lysis Kinetics and Enhances Efficacy.

Drug-resistant bacterial pathogens are a serious threat to global health, and antibacterial lysins are at the forefront of innovative treatments for these life-threatening infections. While lysins' general mechanism of action is well understood, the design principles that might enable engineering of performance-enhanced variants are still being formulated. Here, we report a detailed analysis of molecular determinants underlying the efficacy of lysostaphin, a canonical anti-MRSA (methicillin-resistant ) lysin. Systematic analysis of bacterial binding, growth inhibition, lysis kinetics, and therapeutic efficacy revealed that binding affinity, and not inherent catalytic firepower, is the dominant driver of lysostaphin efficacy. This insight enabled electrostatic affinity tuning of lysostaphin to produce a single point mutant that manifested dramatically enhanced processivity and lysis kinetics and trended toward improved efficacy. More generally, these studies provide important insights into the complex relationships between lysin electrostatics, bacterial targeting, cell lysis efficiency, and efficacy. The lessons learned may enable engineering of other high-performance antibacterial biocatalysts.