Glyceraldehyde3-phosphate dehydrogenase (GAPDH) is a multifunctional protein involved in celldeath processes frequently associated with oxidative/nitrosativo stress.S-nitrosylation of GAPDH facilitates its binding to the E3-ubiquitin-ligase Siah1,which has a nuclear localization signal that promotes the entrance of the proteincomplex to the nucleus causing apoptosis. GOSPEL (GAPDH's Competitor O f Siah1Protein Enhances Life) protein interacts with GADPH and interferes with thebinding between GAPDH and Siah1, inhibiting their apoptotic effect.
Oxidative/nitrosativestress also induces the aggregation of GAPDH in vitro, which is in accordance with the presence of the enzyme ininsoluble aggregates found in some neurodegenerative diseases.
Evidenceprovided by our laboratory (1) indicates that in the presence of nitric oxide(NO) GOSPEL co-aggregates with GAPDH increasing its aggregation rate. GAPDHCys152 plays an essential role in this process since their S-nitrosylation initiatesthe oxidative modification that triggers the formation of disulfide-bondedaggregates. Both GAPDH aggregation and GAPDH-GOSPEL co-aggregation were inhibitedby NAD+ .
Here we present preliminary circular dichroismstudies indicating that NAD+ inhibitsthe conformational changes induced by the NO donor NOR3. We also report theX-ray structure of GAPDH treated with NOR3 in the presence of NAD+. In addition to the NAD+ densitythe difference map exhibits a positive density connected to the SH of Cys152that could only be attributed to NO. These results suggest that NAD+ couldbe inhibiting the NOR3-induced aggregation by stabilizing NO-GAPDH conformation