The 'assembly-promoting sequence region' of microtubule-associated protein 4 failed to promote microtubule assembly

In order to study the function of the bovine MAP4 microtubule-binding domain (the assembly-promoting (AP) sequence region), a fragment corresponding to the AP sequence region was prepared using an Escherichia coli expression system. When the fragment was mixed with purified tubulin at 37 degrees C, the fragment caused a time- and dose-dependent turbidity increase, and the fragment bound to tubulin. However, the products were cold-stable, and amorphous aggregates were observed by electron microscopy. Using axonemes as the seeds for microtubule assembly, the microtubule-elongating activity of the fragment was examined. A dose-dependent turbidity increase of the sample was observed, and electron microscopic observation revealed that microtubules were dose-dependently elongated from the axonemes. Consequently, the AP sequence region does not nucleate microtubules, but elongates them.