We seek to understand how group A Streptococcus defeats one of the primary human defenses against infection, the cationic antimicrobial peptide LL-37. Group A Streptococcus (GAS, S. pyogenes) is a leading cause of global morbidity and mortality. With an estimated >500,000 deaths annually, GAS ranks among the top 10 causes of mortality from infectious disease. GAS is responsible for acute invasive diseases, such as streptococcal toxic shock syndrome (STSS) and necrotizing fasciitis (NF), which have alarmingly high mortality rates (as high as 80% in some cases). The leading cause of acute invasive GAS disease worldwide for more than 30 years has been a globally disseminated subclone of the GAS M1T1 serotype. A critical virulence determinant of the GAS M1T1 serotype strain is the M1 protein. We have shown recently in LaRock et al. (Cell Host & Microbe 2015) that one of the critical functions of the M1 protein is detoxification of the human cathelicidin LL-37, a cationic antimicrobial peptide. LL-37 forms amphipathic α-helices, and functions, like other amphipathic α-helical cationic antimicrobial peptides, by inserting into bacterial plasma membranes and causing lysis. The M1 protein enables GAS M1 serotype strains to resist killing by LL-37, and to resist killing within neutrophil extracellular traps (NETs), where LL-37 is abundant. The M1 protein does this by sequestering LL-37 into a ‘protein trap’, which prevents LL-37 from interacting with its target of action, the bacterial membrane. Importantly, we have found that the virulence contribution of the M1 protein in a murine subcutaneous wound infection model is almost entirely attributable to its interaction with the murine ortholog of LL-37, CRAMP. How the M1 protein binds and detoxifies LL-37 is unknown, and is the focus of this proposal. Our specific aims are to (1) Identify the M1 amino acids responsible for binding and detoxifying LL-37, and (2) Elucidate M1-cathelicidin interactions at the atomic level. This second aim is a high-risk, high-reward aim that is in perfect alignment with the goals of the R21 mechanism. Knowledge gained from this study will be applicable to the design of strategies aimed at restoring the antimicrobial power of LL-37 on GAS M1 strains. This is a significant goal as the GAS M1 strain is the predominant cause of invasive GAS disease in the US, surpassing the next most frequent GAS M type strain by more than 3-fold, and as noted above, an M1T1 serotype strain is the predominant cause of invasive GAS disease worldwide. Furthermore, it seems highly unlikely that the M1 protein would be the only M protein to bind and detoxify LL-37. A number of invasive GAS strains are resistant to the action of LL-37, and indeed we have direct evidence that other M proteins bind and detoxify LL-37. Thus, this project will not only have direct impact on combatting the widespread and medically significant GAS M1 strain, but will also likely have broader implications.