The SPRY domain-containing SOCS box protein 2 (SPSB2) interacts with inducible nitric oxide synthase (iNOS), a critical effector of the innate host response that is required for the intracellular killing of pathogens such as Mycobacterium tuberculosis and Leishmania major.1,2 The SPSB2-iNOS interaction results in proteosomal degradation of iNOS. Consistent with this, SPSB2-deficient macrophages have significantly elevated levels of both iNOS and nitric oxide (NO). This suggests that inhibitors of SPSB2-iNOS interactions may prolong the lifetime of iNOS and therefore NO production, and thereby enhance the killing of persistent pathogens.
In this study, cyclic peptides were designed and synthesized based on the key binding motif of the SPSB2-iNOS interactions i.e. the DINNN peptide sequence of iNOS.3 When the cyclic peptides were added to SPSB2, significant chemical shift perturbations were observed in the 1H-15N HSQC spectrum of SPSB2. Interestingly, the same resonances in the HSQC spectrum of SPSB2 were perturbed when the linear DINNN peptide was added, indicating that the cyclic peptides bound to the same iNOS binding site. When surface Plasmon resonance (SPR) experiments were carried out to determine the binding kinetics and affinities of the cyclic peptides, low nanomolar binding affinities were observed. Our results suggest that these cyclic peptides are potent ligands for SPSB2, and therefore inhibitors of the SPSB2-iNOS interaction with the potential to be a new class of anti-infectives. Further optimization of these peptides is currently in progress.