Involvement of the carboxyl-terminal domain of tubulin in the regulation of its assembly.
Limited proteolysis of phosphocellulose-purified tubulin with subtilisin resulted in cleavage of both alpha and beta tubulin subunits, with the formation of two major fragments (S alpha, and S beta, 48 kDa) and a small peptide (4 kDa) containing the carboxyl-terminal region of tubulin. Interestingly, tubulin cleaved under the present conditions showed an increased ability to assemble into large polymers in the absence of MAPs and under conditions that do not promote assembly of undigested tubulin--i.e., low magnesium concentrations and the absence of taxol and polyalcohols. The critical concentrations for the subtilisin-cleaved tubulin assembly was similar to that of MAPs-promoted tubulin assembly. Assembly product from subtilisin-cleaved tubulin consisted mainly of protofilament bundles, hooked polymer, and open tubules, structures showing equatorial and longitudinal spacings of 50 and 40 A, respectively. The existence of junctions between polymer walls indicates that the carboxyl-terminal removal facilitates polymer-polymer interactions. These results, together with previous studies on the involvement of the carboxyl-terminal domain of tubulin in its interaction with MAP-2, suggest a regulatory role for this domain in tubulin assembly. Thus, in general terms the tubulin molecule can be analyzed as a protein containing two essential domains with functional significance, one domain playing a major role in self-association and the other (the carboxyl-terminal moiety) playing a regulatory role in modulating the interactions responsible for self-association.
D000818:Animals; D001921:Brain / Q000378:metabolism*; D007700:Kinetics; D046911:Macromolecular Substances; D008854:Microscopy, Electron; D008868:Microtubule Proteins / Q000378:metabolism; D008970:Molecular Weight; D010446:Peptide Fragments / Q000032:analysis; D013381:Subtilisins; D013552:Swine; D014404:Tubulin / Q000302:isolation & purification / Q000378:metabolism*
D046911:Macromolecular Substances; D008868:Microtubule Proteins; D010446:Peptide Fragments; D014404:Tubulin; D013381:Subtilisins