Analysis of the Inhibitory Effects of Suramin on Inducible and Neuronal NOS
Carroll, Andrew J.
MetadataShow full item record
Nitric oxide synthase has gained momentum and significant interest in the scientific community due to its vast importance in the human body. Nitric oxide synthase (NOS) is a significant biological enzyme whose product (-NO) is both needed physiologically, and has been linked in pathological conditions such as Alzheimer's Disease. Suramin is a polysulfonated naphthylurea hexasodium salt that has already been in extensive use clinically as it is effective in treating diseases such as onchocerciasis and prostate cancer. The present study was undertaken to explore the ability of Suramin to differentiate between isoforms of nitric oxide synthase. E. coli expressed NOS was obtained, purified and activity and inhibitory effects of Suramin were determined by hemoglobin assays. The extent of purification was assessed by SDS-PAGE analysis. The extent of inhibition was determined by Dixon plots. Suramin was shown to inhibit both nNOS (Km ~ 3 uM) & iNOS (Km - 12 uM), however Suramin had a greater effect on iNOS. Suramin was able to inhibit the neuronal isoform (Ki = 75.11 uM) however it proved to be a more potent inhibitor of the inducible isoform (Ki = 2 uM). The ratio of Ki values indicates an approximate 37 fold selectivity of the inducible over the neuronal isoforms. Dixon analysis suggests that Suramin acts as a non-competitive parabolic inhibitor of the neuronal isoform, and as an uncompetitive inhibitor ofthe inducible isoform. Western analysis also suggests that Suramin binds by interacting at the calmodulin binding site of iNOS. Since Suramin has been demonstrated to select among calmodulin binding proteins, it is a potential molecule to study differences within NOS.