Structural basis for the regulation of tubulin by vinblastine

Abstract

Vinblastine is one of several tubulin-targeting Vinca alkaloids that have been responsible for many chemotherapeutic successes since their introduction in the clinic as antitumour drugs. In contrast with the two other classes of small tubulin-binding molecules (Taxol and colchicine), the binding site of vinblastine is largely unknown and the molecular mechanism of this drug has remained elusive. Here we report the X-ray structure of vinblastine bound to tubulin in a complex with the RB3 protein stathmin-like domain (RB3-SLD). Vinblastine introduces a wedge at the interface of two tubulin molecules and thus interferes with tubulin assembly. Together with electron microscopical and biochemical data, the structure explains vinblastine-induced tubulin self-association into spiral aggregates at the expense of microtubule growth. It also shows that vinblastine and the amino-terminal part of RB3-SLD binding sites share a hydrophobic groove on the alpha-tubulin surface that is located at an intermolecular contact in microtubules. This is an attractive target for drugs designed to perturb microtubule dynamics by interfacial interference, for which tubulin seems ideally suited because of its propensity to self-associate.

Stathmin/Op18 is a novel mediator of vinblastine activity

“…In the presence of stathmin, we observed a 50-fold increase in the VLB-tubulin binding constant. The fact that VLB increases stathmin binding on tubulin, and vice versa, is in good agreement with the X-ray structure of the RB3-tubulin complex which revealed that both stathmin and VLB binding induce similar conformational consequences, curving consecutive tubulin dimers [13].…”

“…However, despite a variety of data from cell lines, the molecular basis of this process is not known. Recent crystallographic data suggest that both VLB and stathmin might act together, since they can bind simultaneously on free tubulin and each results in the curving of tubulin dimer filaments [13]. In this study, we present the first direct evidence of the functional synergy between endogenous stathmin and the antimitotic drug VLB.…”

“…2. tubulin is the same in T2S as in VLB-induced oligomers [13], this difference in the DC p observed in the two binding processes (B2 and A1) can be explained by the positive contribution to DC p from the burying of inter-tubulin interface hydrophobic surfaces [26] during VLB-induced tubulin oligomerization. These values of DC p enabled us to calculate DH = 2 kcal mol À1 for VLB binding to tubulin and DH = À20 kcal mol À1 for VLB binding to T2S at 37°C.…”

Amino acid substitutions at proline 220 of β-tubulin confer resistance to paclitaxel and colcemid

“…Vinca alkaloids, such as vinblastine, vincristine, and vinorelbine, occupy a third binding site that is predominantly in the a-tubulin subunit and sits at the interface between adjacent heterodimers in the protofilament (16). Occupation of this site destabilizes microtubules by inducing a tubulin conformation that favors spiral curvature in the protofilaments (16,17).…”

Developmental Biology of Neoplastic Growth

“…However, the binding of dolastatin 15 was inhibited not only by cryptophycin 1 and dolastatin 10, but also by halichondrin B and vinca alkaloids. The vinca domain seems to be located in β-tubulin at the inter-dimer interface between α-tubulin and β-tubulin subunits in each protofiber (Gigant et al 2005), suggesting that these toxins do not bind to the vinca domain itself, but the binding site(s) for these toxins consist(s) of a series of overlapping domains on the surface of tubulin (Cruz-Monserrate et al 2003). This is consistent with the finding that hemiasterlin binds to helix H10 on α-tubulin which interacts across the interdimer interface with helix H6 of β-tubulin (Nunes et al 2005).…”