Detection of Disulfide-Linked Peptides by Mass Spectrometry
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Mass spectrometry is playing a rapidly increasing role in protein chemistry and sequencing (see Chapter 91) and is particularly useful in determining sites of co- and posttranslational modification (1,2), and application in locating disulfide bonds is no exception. This technique can of course readily analyze peptide mixtures. Therefore, it is not always necessary to isolate the constituent peptides. A combination of microsequencing and mass spectrometry techniques is now commonly employed for complete covalent structure determination. On-line electrospray mass spectrometry (ESMS) coupled to capillary electrophoresis or high-performance liquid chromatography (HPLC) has proven particularly valuable in the identification of modified peptides (3). Recent developments in ESMS sources permit on-line microbore HPLC using matrices, such as 10-µm Poros resins slurry-packed into columns <0.25 mm in diameter. Polypeptides can be separated on gradients of 5–75% acetonitrile over 2 min in formic or acetic acid (0.1%) (4).
KeywordsDisulfide Bond Cyanogen Bromide Mass Spectrometry Technique Intramolecular Disulfide Bond Performic Acid
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