Polyamine Sharing between Tubulin Dimers Favours Microtubule Nucleation and Elongation via Facilitated Diffusion
Figure 10
Effect of spermidine on microtubule elongation.
(A) Pseudo-first order rate constant of elongation, Kobs, versus tubulin concentration in buffer M. 100 µM spermidine significantly increases the elongation rates whatever tubulin concentration. Inset: Log-plot of 1−I(t)/I(∞) versus time for 30 µM tubulin. The slope of this curve is −Kobs. The elongations rate is about three times higher with spermidine (except for 40 µM tubulin). (B) A microtubule solution was prepared by incubating 30 µM tubulin at 37°c for 1h in 50 mM MES-KOH pH 6.8, 1 mM EGTA, 2 mM MgCl2, 20% glycerol, 1 mM GTP. At the end of the incubation, 50 µl of a solution containing 30 µM of free tubulin dimers without or with 100 µM spermidine was then added to 50 µl of the microtubule solution. The sudden increase of the free tubulin concentration allows one to observe elongation of the preformed microtubule via light scattering. It turns out that the presence of 100 µM spermidine leads to a significant increase of the elongation rate as indicated by the slope of the assembly curve, which is 2.7 times higher with 100 µM spermidine (see dotted lines). The plateau value which is more rapidly reached in the presence of polyamines also evidences a facilitated elongation.