Practical Protein Chemistry - A. Darbre 1989

Disulfide bonds
Special methods
Diagonal mapping

The diagonal map method was specifically designed for the analysis of cystine-containing Peptides [4, 6]. Various modifications of this method are known, but its original version—paper Electrophoresis—is most frequently used. The enzymatic hydrolyzate of the protein under study is separated by paper electrophoresis on Whatman 3MM paper (400×400 mm). The electrophoretogram is then dried and exposed to performic acid vapors for 2 h to oxidize the S—S bonds. After drying in a vacuum desiccator over NaOH, electrophoresis is carried out under analogous conditions, but in a direction perpendicular to the initial one. As a result, all peptides that were unmodified during oxidation lie along the diagonal on the electrophoretogram at a 45° angle. Cystine-containing peptides that underwent intermediate oxidation form pairs of fragments located off the diagonal. These zones are eluted, and the resulting peptides are analyzed using standard Methods. To isolate cystine-containing peptides after the first electrophoresis, a control strip can be cut out, treated with performic acid vapors, sewn onto another sheet of paper, and subjected to a second electrophoresis. By comparing the resulting electrophoretogram with the initial one, one can localize the zones corresponding to the cystine-containing peptides, cut them out, and elute the intact material for subsequent operations. Known modifications of this method include the preparation of various Cysteine derivatives, such as S-aminoethylcysteine [33].