A quantitative analysis of tubulin-colchicine binding to microtubules.

The binding of the tubulin-colchicine complex to microtubules has been studied in conditions where copolymerisation, as observed by Sternlicht and Ringel [J. Biol. Chem. 254, 10540-10550 (1979)], is negligible. The binding is shown to be rapid and reversible, in contrast with assumptions found in the literature. The binding constant was determined by a quantitative analysis of the concentration dependence of growth inhibition. The binding constant is of the same order of magnitude as the equilibrium constant of growth for tubulin. The temperature dependence was also studied and the Van't Hoff plot was found to be biphasic with an exothermic part at temperatures lower than 30 degrees C. A molecular model is presented for the interpretation of the copolymerisation observed by Strenlicht and Ringel. When a large excess of colchicine was added to microtubules, only a small phase of dissociation was observed. The new end state was metastable, as further dissociation could not be reversed. This is in conflict with a pure head-to-tail polymerisation mechanism, but in agreement with the recent observations of Bergen and Borisy [J. Cell Biol. 84, 141-150 (1980)], that head-to-tail polymerisation contributes only a small fraction of the total polymerisation.