Figure 2: GLB-12 is a hexacoordinated globin with redox properties and is capable of producing O2.−.
From: A redox signalling globin is essential for reproduction in Caenorhabditis elegans
(a) UV–vis spectra of different GLB-12 forms, with their absorption maxima. The absorption spectrum of purified GLB-12 showed that it is spontaneously oxidized to the ferric state (Fe3+) upon exposure to air. Reducing GLB-12 (Fe2+) resulted in an absorption spectrum that is typical for a hexacoordinated haem iron. Absorption spectra of CN- and CO-bound GLB-12 could be obtained after exposure of GLB-12 to an excess of KCN or CO. (b) The current potential behaviour of a gelatin B (GelB) and a GelB/GLB-12 electrode, with the reduction (Epc) and oxidation peak (Epa), showing that GLB-12 is reversibly oxidized and reduced with a reduction potential of −0.244 V (versus SCE). The reversible oxidation/reduction of GLB-12 by cyclic voltammetry is illustrated in the scheme. (c) The current potential behaviour of a gelatin B electrode containing GLB-12 and CCP (cytochrome C peroxidase), with and without H2O2. The arrow points to the increase of the reduction peak when H2O2 is added, showing that GLB-12 becomes fully reduced by electron transfer to CCP, making CCP capable of reducing H2O2. This reaction is illustrated in the scheme. (d) GLB-12 is capable of producing O2.−. (+) indicates the presence, (−) the absence of glucose-6-phosphate and GLB-12. (e) Comparison of in vitro superoxide production by GLB-12, cytochrome C, myoglobin and cytochrome P450 (Cyt P450). Myoglobin and cytochrome c, which have a reduction potential higher than the O2/O2.− couple64,65 and therefore should not be able to produce O2.−, did not lead to an increase in luminescence. Cytochrome P450, which has a reduction potential lower than the O2/O2.− couple and can produce high levels of O2.− (ref. 66), led to a large increase in luminescence. GLB-12 led to a moderate increase in luminescence, indicating that it produced moderate levels of O2.−. (f) The antioxidants ascorbic acid and N-acetylcysteine (NAC) enhance the glb-12 RNAi reduction in fecundity (n=4). Percentages show control RNAi compared with glb-12 RNAi. Data are represented as mean±s.e.m.; *P<0.05 (two-sided Student t-test).
