Autonitrosylation of di-ferrous YtfE by
nitrite. (A) UV–visible
absorbance spectra of di-ferrous YtfE (black line) and nitrite-treated
di-ferrous YtfE (red line) post desalting. Air-exposed (mixed-valent/di-ferric)
YtfE is shown for comparison (green line). Inset: the difference spectrum
between mixed-valent and nitrite-treated YtfE reveals a band at 397
nm, indicative of iron-nitrosyl species. (B) EPR spectra of as-isolated
di-ferrous YtfE, pre- and post-treatment with nitrite. Signals arising
from MNIC (g = 3.95), DNIC (g =
2.03), and mixed-valent YtfE (g = 1.96, 1.91, and
1.88) are indicated. Note that the EPR spectrum of air-exposed YtfE
is shown in Figure 3D. The native mass spectrometry of di-ferrous YtfE treated with (C)
nitrite or (D) ethyl nitrite. Nitrosylated YtfE is observed after
nitrite, but not ethyl nitrite, treatment. An ethyl nitrite adduct
(26,230 Da) was the major species in the latter. A minor peak due
to an acetate adduct of the ethyl nitrite adduct was also observed
at 26,289 Da. On the low mass side, an oxygen atom (+16 Da) adduct
of YtfE was observed at 26,169 Da. The lower spectrum in panel (D)
was obtained through in-source collision (isCID), which resulted in
the breakdown of ethyl nitrite to give the nitrosylated adduct, along
with some unknown lower mass species.