Inducible nitric-oxide synthase (iNOS) plays a pivotal role in combatting leishmaniasis and tuberculosis through nitric oxide (NO) production mechanism. iNOS is negatively regulated by ubiquitination and proteasomal degradation. iNOS contains a highly conserved DINNN motif in its N-terminal region that strongly interacts with the SPRY-domain containing SOCS box proteins (SPSBs). Upon binding, SPSBs recruits an E3 ligase ubiquitin ligase complex to polyubiquitinate iNOS, targeting it for proteasomal degradation. By blocking this key interaction between SPSB2 and iNOS, the intracellular lifetime of iNOS is prolonged, which may enhance the killing of infectious pathogens such as bacteria and Leishmania major parasites.1-3

Using NMR-based approaches such as Saturation Transfer Difference (STD-NMR) and Carr-Purcell-Meiboom-Gill (CPMG), we have successfully screened the MIPS fragment library of 1140 compounds and found 12 fragments that bind to SPSB2. Using the competition STD-NMR and CPMG experiments, we confirmed that these 12 fragments bind at the SPSB2-iNOS interaction site. A series of analogues relating to these fragment hits has been tested by NMR and surface Plasmon resonance. Further elaboration of these analogues is currently underway to generate more potent compounds that may serve as leads in the development of novel anti-infective agents.

1. Kuang et al. (2010) J. Cell. Biol. 190, 129-141.
2. Kuang et al. (2009) J. Mol. Biol. 386, 662-674.
3. Lewis et al. (2011) J. Immunol. 187, 3798-3805.