Mapping the Protein—Protein Interface between a Toxin and Its Cognate Antitoxin from the Bacterial Pathogen Streptococcus pyogenes

Protein–protein interactions are ubiquitous and essential for most biological processes. Although new proteomic technologies have generated large catalogs of interacting proteins, considerably less is known about these interactions at the molecular level, information that would aid in predicting protein interactions, designing therapeutics to alter these interactions, and understanding the effects of disease-producing mutations. Here we describe mapping the interacting surfaces of the bacterial toxin SPN (Streptococcus pyogenes NAD+ hydrolase) in complex with its antitoxin IFS (immunity factor for SPN) by using hydrogen–deuterium amide exchange and electrospray ionization mass spectrometry. This approach affords data in a relatively short time for small amounts of protein, typically 5–7 pmol per analysis. The results show a good correspondence with a recently determined crystal structure of the IFS–SPN complex but additionally provide strong evidence for a folding transition of the IFS protein that accomp...