Retracing the origin and evolution of a cryptic antimicrobial peptide within mammalian lactoferrin

Antimicrobial peptides (AMPs) constitute key components of innate immunity across the tree of life. Canonical AMPs are typically translated as small proteins and secreted from host cells to act against microbes. However, cryptic AMP-like domains are also embedded within diverse proteins not classically associated with antimicrobial function. How such embedded AMPs first emerge and diversify remains unclear. Here we retrace the origin and evolution of the abundant mammalian protein lactoferrin and its embedded AMP, lactoferricin. By resurrecting extinct lactoferrin ancestors dating back to the earliest mammals, we identify an enrichment of cationic and hydrophobic amino acids in the lactoferricin domain over time. These changes enabled ancient lactoferricin to first rupture bacterial membranes, an activity that was later enhanced in extant mammals conferring potent bactericidal activity. In addition, we find that natural selection within the lactoferricin domain has continued to modulate antimicrobial activity on recent evolutionary timescales. In particular, we pinpoint a single rapidly evolving site in lactoferricin among great apes that significantly enhances antimicrobial potency against major pathogenic bacteria. Together, our study illustrates how novel immune protein functions can arise, evolve, and diversify to strengthen host defense against microbial pathogens.