Identification of an artifact in the mass spectrometry of proteins derivatized with iodoacetamide

✍ Scribed by Lapko, Veniamin N.; Smith, David L.; Smith, Jean B.

• Publisher: John Wiley and Sons
• Year: 2000
• Tongue: English
• Weight: 78 KB
• Volume: 35
• Category: Article
• ISSN: 1076-5174
• DOI: 10.1002/(sici)1096-9888(200004)35:4<572::aid-jms971>3.0.co;2-2

✦ Synopsis

Derivatization of cysteinyl residues is often used to prevent the formation of disulfide bonds during protein isolation and analysis. The most commonly used reagents are iodoacetic acid and iodoacetamide, which increase the molecular mass of the protein by 58 or 57 Da, respectively, for each derivatized cysteine. A possible side reaction is derivatization of methionine. In our analysis of derivatized human lens aA-crystallins, we found an apparent molecular mass 48 Da lower than the mass expected for aA-crystallin with the cysteines carboxyamidomethylated. Analysis of a tryptic digest of this protein showed that both cysteines and one methionine had been derivatized. Peaks indicating a molecular mass 48 Da less than expected for the protein with only cysteines derivatized were attributed to fragmentation of the derivatized methionine through collisioninduced dissociation in the electrospray ionization source. An awareness of this artifact is important to investigators searching for proteins and their modified forms in complex mixtures.