The molecular mechanism of interaction of Et3Pb+ with tubulin

Triethyllead ion (Et3Pb+) was found to interact with 2 out of 18 thiol groups present in tubulin dimers. Specificity of the interaction was shown by the high affinity of Et3Pb+ to tubulin, by the fact that the 16 residual thiol groups in tubulin remained unaffected, and by the observation that other proteins with exposed thiol groups, e.g., actin, did not react with Et3Pb+. After complexation of the two thiol groups, tubulin in vitro had lost its capability for microtubule assembly. Likewise, polymerized tubulin disassembled on addition of the lead compound

In vitro formation of different tubulin polymers from purified tubulin of ehrlich ascites tumor cells.

AbstractPreparations of cycled tubulin from Ehrlich ascites tumor cells contain several acessory proteins; once or twice cycled microtubule preparations are usually composed of fibers 10 nm in diameter, but lack vimentin. Highly purified tubulin consists of α- and β-tubulin and a minor component which was identified by peptide mapping as a second β-chain. This pure tubulin is able to form in vitro at low concentrations (1 mg protein/ml) fibers of about 10 nm width, and at higher concentrations (3.5 mg protein/ml) normal microtubules

High cytotoxicity and membrane permeability of Et3Pb+ in mammalian and plant cells

Cells of mammalian origin as well as those of higher plants appear to be very sensitive to triethyllead ion (Et3Pb+). Neuroblastoma cells kept in the presence of 1 μM Et3Pb+ lost their viability within 6 h. Growth of suspension culture cells of soybean (G. max (L.)Merr.) was inhibited by 1 μM Et3Pb+, and finally the cells died. Morphologically, Et3Pb+ caused the complete breakdown of microtubular structures in neuroblastoma cells; thus microtubules appeared to be the main target for the toxin. While in a previous study the effect of Et3Pb+ on microtubules has been well documented at concentrations of 50–200 μM 1, the present study demonstrates that the formation of microtubules from pig brain tubulin is disturbed at concentrations of Et3Pb+ as low as 0.5 to 1 μM . We conclude from these data that Et3Pb+ freely permeates the plasma membranes of mammalian as well as plant cells

The interaction of triethyl lead with tubulin and microtubules

The impact of triethyl lead chloride was studied on: (i) the in vitro assembly and disassembly of microtubules from porcine brain by turbidometry and electron microscopy, (ii) the microtubule system of living mammalian cells using immunofluorescence microscopy, (iii) cell motility and chemotaxis employing the methods of phagokinetic track formation and the Boyden chamber assay, respectively, and (iv) thiol groups of the protein tubulin by their titration in the presence and absence of the organic lead compound. Triethyl lead chloride inhibited microtubule assembly and depolymerized preformed microtubules in vitro and in living cells. Random motility of cells was not markedly inhibited by triethyl lead chloride, whereas chemotaxis (directed cellular movement) was strongly inhibited. Triethyl lead chloride was found to interact with 2 thiol groups of the tubulin dimer. The interaction of triethyl lead chloride with the tubulin/microtubule system in vivo likely causes aneuploidy and is at least partly responsible for the cytotoxicity of the drug