a-Tubulin is biosynthesized with a tyrosine at its C-terminus which can be removed by tyrosine carboxypeptidase and re-incorporated by tubulin tyrosine ligase. We studied the capability of the ligase to incorporate tyrosine analogues into tubulin. Azatyrosine induces the reversion of cancerous phenotype and can be incorporated into the tubulin?s C-terminus as well as into proteins via de novo synthesis. Which of these two mechanisms is responsible of this effect remains unclear. The introduction of a nitro group into the position 3 of the phenolic ring of tyrosine avoids its incorporation into proteins via de novo synthesis, but not into tubulin?s C-terminus. Therefore, 3-Nitroazatyrosine was synthesized, purified by TLC and characterized by UV-Visible, IR, mass and RMN-1H spectroscopy. It was found that 3-nitroazatyrosine cannot be incorporated into proteins via de novo synthesis nor into tubulin?s C-terminus. No effect was found in cellular proliferation. We also studied if the ligase was able to incorporate other tyrosine analogues. Melphalan could not be incorporated; Thienylalanine and p-aminophenylalanine were incorporated with low affinity, and m-fluoro-tyrosine was incorporated very efficiently in vitro. m-F-tyrosine stopped proliferation of C6 cells and changed their morphology. An antibody against the C-terminal m-F-Tyr residue was developed.
