We have identified conditions that allow metal-catalyzed oxidation (MCO) reactions
and mass spectrometry (MS) to correctly identify binding sites of first-row transition
metal ions to model peptides. This work extends the applicability of the MCO/MS method
to metals other than Cu(II). When the appropriate reducing agent (ascorbate, 10 mM)
and oxidizing agent concentrations (1 mM persulfate, atmospheric O2, or both) are
used, metal-bound amino acids can be sufficiently and specifically oxidized for clear
identification by MS. The MCO reactions with Mn(II), Fe(II), Co(II), and Ni(II) occur
to lesser extents than with Cu(II), but oxidation is still extensive enough to allow
easy identification of the metal-bound residues. With the exception of aspartic acid,
the known metal-binding amino acids of angiotensin I and bacitracin A are oxidized,
while no oxidation is observed at nonbinding residues. Failure to oxidize aspartic
acid is likely due to the relatively slow reactivity of its carboxylic acid side chain
with reactive oxygen species, suggesting that the current MCO/MS protocol is transparent
to such acidic residues. Overall, this study indicates that, just as is possible for
Cu(II), the MCO/MS method should be suitable for determining the Mn(II)-, Fe(II)-,
Co(II)-, and Ni(II)-binding sites of metalloproteins.