Induction profiles and properties of a novel stress-induced peroxidase in Neurospora crassa

Exposure of Neurospora crassa cells to heat shock and oxidative stress results in the synthesis of several stressrelated proteins, including a peroxidase. Northern blot analysis of total RNA revealed a heat-inducible (HI)-peroxidase transcript of ~10kb, induced in response to heat shock and oxidative stress. The HI peroxidase was isolated from heat-shocked mycelium and purified to near homogeneity, and its properties were examined. Chromatography in size-exclusion matrices yielded an apparent molecular mass of ~116kDa for the native enzyme, whereas the estimate obtained by SDS-PAGE was 90–95kDa. Studies of substrate saturation kinetics were conducted using the purified enzyme with ABTS [2,2’-azino-bis (3- ethylbenzthiazole-6-sulfonic acid)] and H2O2 as substrates. The experimentally estimated Km, Vmax, and Kcat values for ABTS were ~36μM, 5200nmolmg−1, and 8s−1, respectively, and those for H2O2 were 44μM, 6640nmolmg−1, and 10s−1. O-dianisidine was a substrate for this enzyme, but guaiacol was not. HI peroxidase was found to be a glycoprotein, stable at temperatures up to 60°C.