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In mass spectrometry‐based peptide sequencing, electron transfer dissociation (ETD) and electron capture dissociation (ECD) have become well‐established fragmentation methods complementary to collision‐induced dissociation. The dominant fragmentation pathways during ETD and ECD primarily involve the formation of c‐ and z•‐type ions by cleavage of the peptide backbone at the N─Cα bond, although neutral losses from amino acid side chains have also been observed. Residue‐specific neutral side chain losses provide useful information when conducting database searching and de novo sequencing. Here, we use a combination of infrared ion spectroscopy and quantum‐chemical calculations to assign the structures of two ETD‐generated w‐type fragment ions. These ions are spontaneously formed from ETD‐generated z•‐type fragments by neutral loss of 33 Da in peptides containing a cysteine residue. Analysis of the infrared ion spectra confirms that these z•‐ions expel a thiol radical (SH•) and that a vinyl C═C group is formed at the cleavage site. z•‐type fragments containing a Cys residue but not at the cleavage site do not spontaneously expel a thiol radical, but only upon additional collisional activation after ETD.