The effects of indole alkaloids and related compounds on the properties of brain microtubular proteins

Abstract

Microtubular subunit protein (tubulin) was isolated from foetal rabbit brain extracts by chromatography on DEAE-Sephadex or by in vitro polymerisation procedures. The effects of indole alkaloids and related compounds on the colchicine and nucleotide binding properties of tubulin were investigated using sensitive radiochemical assay procedures developed for this purpose. It was shown that in contrast to the stabilising effects of Vinca alkaloids (vincristine and vinblastine), a number of structurally related drugs containing methoxyindole derivatives (e.g. reserpine, melatonin, harmine) inhibited the binding of colchicine to the protein. However, both groups of indole alkaloids markedly enhanced the binding of guanosine nucleotides to tubulin. During the course of this work, the specificity of nucleotide binding sites present in isolated microtubular protein preparations was also investigated.

The effects of indole derivatives on the in vitro assembly of brain microtubules were studied by electron microscopy: various Vinca alkaloids as well as colchicine caused either partial or complete block of microtubule assembly, while melatonin promoted the appearance of 'twisted' microtubules. However, the reassembly of microtubules in the presence of reserpine appeared to be normal. Additional experiments are reported concerning the binding of [3H] vincristine to isolated tubulin and the subcellular distribution of vincristine binding receptors in brain homogenates. A short collaborative study on the phosphorylation of microtubular proteins in vitro and in situ is also reported.