TUBULIN FOLDING IS ALTERED BY MUTATIONS IN A PUTATIVE GTP-BINDING MOTIF

Tubulins contain a glycine-rich loop, that has been implicated in micr otubule dynamics by means of an intramolecular interaction with the ca rboxy-terminal region. As a further extension of the analysis of the r ole of the carboxyterminal region in tubulin folding we have mutated t he glycine-rich loop of tubulin subunits. An alpha-tubulin point mutan t with a T-150-->G substitution (the corresponding residue present in beta-tubulin) was able to incorporate into dimers and microtubules. On the other hand, four beta-tubulin point mutants, including the (G148) -->T substitution, did not incorporate into dimers, did not release mo nomers, but were able to form C-900 and C-300 complexes (intermediates in the process of tubulin folding). Three other mutants within this r egion (which approximately encompasses residues 137-152) were incapabl e of forming dimers and C-300 complexes but gave rise to the formation of C-900 complexes. These results suggest that tubulin goes through t wo sequential folding states during the folding process, first in asso ciation with TCP1-complexes (C-900) prior to the transfer to C-300 com plexes. It is this second step that for tubulin folding and assembly.