Spectroscopic snapshots of the CmlI catalytic cycle and HPLC analysis of associated products. The reaction began with 600 μM CmlIred (grey trace), to which O2 was added to form P (orange trace). Anaerobic addition of 600 μM NH2-CAM yields Sample F (solid black trace), proposed to be CmlIred-NO-CAM by comparison to independently prepared 600 μM CmlIred-NO-CAM (dotted black trace). The addition of O2 to Sample F yields Sample G (solid blue trace), which nearly overlays with an independently prepared 600 μM decayed P sample (dotted blue trace). Inset: HPLC analysis of products in Sample F (black trace) and Sample G (blue trace). Sample F yields NO-CAM as the primary product and Sample G shows that NO-CAM has been consumed and CAM produced. Buffer: 50 mM Bicine, pH 9, 4 °C