Figure 6.

Kinetics of YtfE-dependent formation of iron-nitrosyl species. (A) Di-ferrous YtfE-mediated formation of iron-nitrosyl species (measured at 360 nm) in response to increasing concentrations of nitrite. (B) Initial rate analysis of the data (black circles) in comparison to equivalent experiments performed with air-exposed (oxidized) YtfE (red circles). Fitting to a simple Michaelis–Menten equation (black lines) gave K_m values for nitrite of 88 and 242 μM for di-ferrous and mixed-valent YtfE, respectively. Error bars indicate standard deviation (n ≥ 4). (C) Plots of A_360 nm showing the effect of Cys30Ala/Cys31Ala YtfE (blue line) and carboxymethylation (red line). The responses of di-ferrous and mixed-valent YtfE in the presence of 1.5 mM KNO₂ are shown for comparison. See Table 1 for full kinetic parameters.