OUTER DOUBLET TUBULIN REASSEMBLY - EVIDENCE FOR OPPOSITE END ASSEMBLY-DISASSEMBLY AT STEADY-STATE AND A DISASSEMBLY END EQUILIBRIUM

[3H]GTP exchangeably bound to outer doublet tubulin becomes nonexchangeable with exogenous GTP upon incorporation of the tubulin into microtubules in vitro. We have used this property to study the mechanism of outer doublet tubulin exchange with reassembled microtubules in vitro. At apparent equilibrium, net addition and loss of tubulin occur at opposite ends of the microtubules. The apparent equilibrium is actually a steady-state summation of two different reactions which occur at the opposite microtubule ends, which results in a unidirectional flux of tubulin from microtubule assembly ends to disassembly ends. The similarity of this behavior with that shown previously for bovine brain microtubules in vitro [Margolis, R. L., & Wilson, L. (1978) Cell 13, 1] supports our earlier contention that neither the doublet structure nor the stability of outer doublet microtubules in situ is determined solely by the tubulin backbone. Further, the intrinsic assembly-disassembly behavior of tubulins from very diverse sources is strongly conserved. Tubulin loss is readily reversible at the disassembly ends of microtubules polymerized from outer doublet tubulin, indicating that these ends are in equilibrium with tubulin in solution. This does not appear to be the case with bovine brain microtubules that have been assembled in vitro. In addition, the kinetics of podophyllotoxin-induced microtubule depolymerization suggest that addition of podophyllotoxin-tubulin complexes may occur at the microtubule diassembly ends under non-steady state conditions. Thus despite the strong conservation of assembly properties of tubulin from stable sea urchin sperm tail outer doublet microtubules and bovine brain, some differences do exist. These differences may reflect differences in the cellular functions of microtubules. © 1979, American Chemical Society. All rights reserved.