A Novel Endogenous Inhibitor of the Secreted Streptococcal NAD-Glycohydrolase

<div><p>The <em>Streptococcus pyogenes</em> NAD-glycohydrolase (SPN) is a toxic enzyme that is introduced into infected host cells by the cytolysin-mediated translocation pathway. However, how <em>S. pyogenes</em> protects itself from the self-toxicity of SPN had been unknown. In this report, we describe immunity factor for SPN (IFS), a novel endogenous inhibitor that is essential for SPN expression. A small protein of 161 amino acids, IFS is localized in the bacterial cytoplasmic compartment. IFS forms a stable complex with SPN at a 1:1 molar ratio and inhibits SPN's NAD-glycohydrolase activity by acting as a competitive inhibitor of its β-NAD⁺ substrate. Mutational studies revealed that the gene for IFS is essential for viability in those <em>S. pyogenes</em> strains that express an NAD-glycohydrolase activity. However, numerous strains contain a truncated allele of <em>ifs</em> that is linked to an NAD-glycohydrolase−deficient variant allele of <em>spn</em>. Of practical concern, IFS allowed the normally toxic SPN to be produced in the heterologous host <em>Escherichia coli</em> to facilitate its purification. To our knowledge, IFS is the first molecularly characterized endogenous inhibitor of a bacterial β-NAD⁺−consuming toxin and may contribute protective functions in the streptococci to afford SPN-mediated pathogenesis.</p>