The protein SPSB2 mediates proteasomal degradation of inducible nitric oxide synthase (iNOS). Inhibitors of the SPSB2-iNOS interaction may prolong the lifetime of iNOS and thereby enhance the killing of persistent pathogens. We have designed cyclic peptides and peptidomimetics containing the key sequence motif mediating the SPSB2-iNOS interaction, which bind to the iNOS binding site on SPSB2 with low nanomolar affinities, as shown by SPR, [1H,15N]-HSQC and/or 19F NMR. An in vitro assay on macrophage cell lysates showed complete inhibition of SPSB2-iNOS interactions by the most potent cyclic peptide. Furthermore, its solution structure closely matched (backbone RMSD 1.2 Å) that of the SPSB2-bound linear DINNN peptide. The designed peptide was also resistant to degradation by the proteases pepsin, trypsin and chymotrypsin, and stable in human plasma. These cyclic peptides and peptidomimetics exemplify a potentially new class of anti-infective agents that acts on the host innate response, thereby avoiding the development of pathogen resistance.