Some chemical properties of carboxymethyl derivatives of amino acids

Tritium-labeled carboxymethyl derivatives of various functional groups in proteins show ready exchange of the tritium label when exposed to standard protein hydrolysis conditions (6 n HCl, 21 h, 110 °C). While N_?-³H]carboxymethyl lysine does not show any tritium exchange, the two N-³H]carboxymethyl histidine derivatives lose their tritium with a half time of about 15 h, and S-³H]carboxymethyl cysteine loses its tritium with a half time of about 1.5 h. The tritium exchange of S-³H]carboxymethyl methionine was so fast that the derivative could not be prepared with any of the tritium label intact. The rate of exchange for this compound was consequently determined by following the disappearance of the methylene NMR signal when S-carboxymethyl methionine was dissolved in D₂O. The half time for the exchange was about 12 min. Mechanisms involving either a sulfonium ion or enolization of the protonated conjugate carboxylic acid appear to give a satisfactory explanation of the relative stability of the different derivatives. The practical use of the differential rate of hydrogen exchange as a means of establishing rapidly and with small quantities of material the site of carboxymethylation in unknown proteins is suggested.