Fig. 1. GC-MS and frequency-selective NMR analyses of N₂-bound Av1*.

(a) GC elution profiles and (b) GC-MS fragmentation patterns of C₂H₄ generated upon turnover with Av1 under D₂/C₂H₂ (grey), Av1 under N₂/D₂/C₂H₂ (blue), and Av1* under D₂/C₂H₂ (brown) in H₂O-based reactions. The C₂H₄ standard is shown as black dashed lines (in a) and black bars (in b). The m/z ratio is indicated for each trace. (c) Frequency-selective ¹H NMR spectra of NH₄⁺ generated from Av1 (solid blue), Av1* prepared with ¹⁴N₂ (solid red) and Av1* prepared with ¹⁵N₂ (solid brown) upon turnover under Ar in the presence of Av2, MgATP and dithionite. Also shown are the ¹H NMR spectra of Av2 control (dashed black) and Av1 control (dashed blue), generated with only Av2 and only Av1, respectively, under Ar in the presence of MgATP and dithionite, as well as a sum of the two, Av1+Av2 control (dashed green). All samples in a-c contained the same amount of Av1, Av1* and/or Av2. Note the minor bumps in the NMR spectra that likely resulted from degraded proteins or interactions of proteins with reagents.