GAPDHcysNO in Bacterial Virulence: Inhibition of Macrophage iNOS Activity
New investigations provide evidence identifying GAPDHcys-NO as a mechanism which regulates iNOS activity. The latter is a heme-containing protein whose activation from its apo-iNOS form is dependent on heme insertion as the rate-limiting activation event. Of note, preliminary studies demonstrated that NO blocked heme insertion not only in iNOS but in a series of cellular proteins (Waheed et al. 2010). GAPDH was identified as a heme-binding protein by affinity chromatography, co-immunoprecipitation in vivo and by in vitro binding studies (Chakravarti et al. 2010). Its role as a major heme transport and transfer protein responsible for its insertion into apo-iNOS was defined by GAPDH knockdown and overexpression analyses. The former diminished heme insertion and reduced iNOS activity while the latter facilitated heme insertion and stimulated iNOS activity.
S-nitrosylation of GAPDH at its active site cysteine forming GAPDHcys-NO abolished GAPDH transfer activity and diminished iNOS activity. Mutation of the active site cysteine abolished both the formation of GAPDHcys-NO and its regulation of iNOS activity. Intriguingly, denitrosylation of GAPDHcys-NO by thioredoxin 1 restored both heme insertion and iNOS activity (Chakravarti and Stuehr 2012).
There is no demonstrable evidence indicating that the effect of GAPDHcys-NO on iNOS activity presents a bacterial pathogenesis mechanism. That being said, the active participation of bacterial NO synthetases in pathogenesis, the rapidity through which bacterially produced NO is involved as an immunoevasive protocol, the significance of membrane GAPDH in bacterial virulence, the significant amount of GAPDH active site modification during oxidative stress, as well as the diverse methods through which bacterial pathogens seek and sequester “free” Fe++ suggest that the examination of this possibility may present a fertile area of investigation.