Practical Protein Chemistry - A. Darbre 1989

Application of electron impact mass spectrometry for determining the amino acid sequence of peptides and proteins
Sample preparation methods

The presence of various functional groups in the protein molecule was the direct reason why it took more than a decade to develop sophisticated Methods for the chemical conversion of Water-soluble polar Peptides of unknown Structure into chloroform-soluble volatile derivatives suitable for electron-impact mass spectrometry. Systematic research into various methods of chemical peptide modification [16–20] led to the creation of a universal method for preparing volatile peptide derivatives for mass spectrometric analysis. The method makes it possible to modify peptides directly in a mixture containing 5–100 nmol of each component [21, 22], regardless of their Amino Acid Sequence. The scheme for such conversion of peptides into N-acetyl-N,O,S-permethyl derivatives is shown in Fig. 19.1. Arginine-containing compounds are usually converted into ornithyl peptides prior to such chemical modification (Fig. 19.2) [22]. To identify arginine-containing peptides in the sample under study, a portion of the mixture is subjected to Amino acid analysis or analytical Electrophoresis followed by staining [23] (Section 8.14.3). The Use of the latter method in the modern sequencing strategy appears to be quite straightforward, as analytical electrophoresis is used to monitor the stages of chromatographic Separation.