Effects of vanadate on the assembly and disassembly of purified tubulin.
Sodium-orthovanadate (100-700 microM) added to purified pig brain microtubule protein (molar ratios 13-90 moles vanadate/mole tubulin) inhibits to a considerable extent the assembly (up to 65%) and the disassembly rates (up to 60%) of microtubules, as determined by turbidimetry. Vanadate added to preformed microtubules did not appreciably alter the turbidity level of the samples, however, the disassembly rates were decreased in the same manner as when vanadate was added prior to polymerization. Microtubule protein kept on ice for 3-6 hours became more susceptible to vanadate than freshly prepared protein. The effect of vanadate was independent of the GTP concentration at which the polymerization assays were performed (0.025 to 1 mM GTP). In the presence of taxol, which increases the rate and extent of microtubule formation, vanadate had no effect on assembly rates. Disassembly was inhibited, however, much less than in the presence of vanadate alone. Electron microscopy and polyacrylamide gel electrophoresis did not reveal differences between microtubules prepared in the presence or in the absence of vanadate. This is consistent with the notion that vanadate does not interfere with the interaction between tubulin and the high-molecular weight microtubule-associated proteins. Apparently vanadate brings about an allosteric change of the microtubule protein(s) resulting in the abnormal polymerization kinetics of tubulin found in our study. The above results may be relevant for studies where the effects of vanadate on intracellular motility are interpreted as being solely due to a specific inhibition of ATPases.
D000818:Animals; D001921:Brain / Q000378:metabolism; D046911:Macromolecular Substances; D008854:Microscopy, Electron; D008869:Microtubule-Associated Proteins / Q000302:isolation & purification / Q000378:metabolism; D008870:Microtubules / Q000187:drug effects / Q000648:ultrastructure*; D009391:Nephelometry and Turbidimetry / Q000379:methods; D013552:Swine; D014404:Tubulin / Q000378:metabolism*; D014638:Vanadates; D014639:Vanadium / Q000494:pharmacology*
D046911:Macromolecular Substances; D008869:Microtubule-Associated Proteins; D014404:Tubulin; D014639:Vanadium; D014638:Vanadates