Residue-specific radical-directed dissociation of whole proteins in the gas phase.

The rapid identification of proteins from biological samples is critical for extracting useful information in proteomics studies. Mass spectrometry is one among the various methods of choice for achieving this task; however, current approaches are limited by a lack of chemical control over proteins in the gas phase. Herein, it is shown that modification of tyrosine to iodo-tyrosine followed by UV photodissociation of the carbon-iodine bond can be used to generate a radial site specifically at the modified residue. The subsequent dissociation of the protein is largely dominated by radical-directed reactions, including dominant backbone fragmentation at the modified tyrosine. If iodination of the protein is carried out under natively folded conditions, the modification and ultimate fragmentation can typically be isolated to a single tyrosine residue. Some secondary backbone cleavage in the immediate vicinity of the modified tyrosine also occurs, especially if proline is present. In the absence of a reactive tyrosine residue, similar chemistry occurs via iodination at histidine. Possible mechanisms which would lead to the observed a-type fragments at tyrosine and the secondary fragments at proline are discussed. A method for using this type of site-specific information to reduce database searching times in proteomics experiments by several orders of magnitude is outlined.