POLYMERIZATION OF TUBULIN IN THE PRESENCE OF COLCHICINE OR PODOPHYLLOTOXIN - FORMATION OF A RIBBON STRUCTURE INDUCED BY GUANYLYL-5'-METHYLENE DIPHOSPHONATE

GTP-dependent in vitro polymerization of rat brain microtubular protein is inhibited to 50% by substoichiometric concentrations of the antimitotic drugs colchicine (0.12 mol/mol of tubulin) and podophyllotoxin (0.14 mol/mol of tubulin). Substitution of pp(CH2)pG† † Abbreviations used: pp(CH2)pG, guanylyl-5′-methylene diphosphonate; MES, [2-(N-morpholino)]-ethane sulfonic acid; SDS, sodium dodecyl sulfate; MAP, high molecular weight microtubule-associated proteins in the nomenclature of Dentler et al. (1975); tau, microtubule-associated polypeptides in the nomenclature of Weingarten et al. (1975). for GTP, however, results in an extensive microtubular protein polymerization at such concentrations. In the presence of pp(CH2)pG, suprastoichiometric concentrations of podophyllotoxin (19 mol/mol of tubulin) are required to inhibit the polymerization process by 50%. Colchicine is very ineffective since 3 × 105 moles/mole of tubulin are required to give a 50% inhibition. Electron microscopical analysis shows that the polymers formed by microtubular protein in the presence of suprastoichiometric concentrations of drugs are not the normal short microtubules typical of pp(CH2)pG-driven polymerization, but are ribbons with three or four protofilaments. The colchicine content of the harvested ribbons has been measured directly and found to be approximately 0.8 moles colchicine/mole of tubulin. Treatment of microtubular protein with substoichiometric concentrations of drugs results in an increase in the number of protofilaments forming the ribbons. Many of the ribbons can close into morphologically normal microtubules when microtubular protein is treated with only 0.05 moles of either colchicine or podophyllotoxin per mole of tubulin. © 1979.