title: Differential effects of magnesium on tubulin-nucleotide interactions.
abstract: Magnesium-depleted 2-(N-morpholino)ethanesulfonate (Mes), glutamate, tubulin and microtubule-associated proteins were prepared and used to study the effects of exogenously added MgCl2 on tubulin-nucleotide interactions in 0.1 M Mes with microtubule-associated proteins and in 1.0 M glutamate. Endogenous levels of Mg2+ in the systems studied were approximately stoichiometric with the tubulin concentrations and largely derived from the tubulin. We examined the effects of added Mg2+ on tubulin polymerization, GDP inhibition of polymerization, binding of GDP and GTP to tubulin, and GTP hydrolysis. Exogenously added Mg2+ had markedly different effects on these reactions. The order of their sensitivity for a requirement for added Mg2+ was as follows: GTP binding greater than GTP hydrolysis greater than polymerization greater than GDP binding. Inhibition of polymerization by GDP varied inversely with the Mg2+ concentration and was greatest in the absence of the cation. These results indicate that GDP and GDP-Mg2+ interact with similar affinity at the exchangeable site, while GTP-Mg2+ has a higher affinity for tubulin than does free GTP. Nevertheless, under appropriate conditions, free GTP can interact sufficiently well with tubulin to permit both nucleation and elongation reactions.
mesh_terms: D000818:Animals; D002021:Buffers; D002417:Cattle; D004492:Edetic Acid / Q000494:pharmacology; D005971:Glutamates / Q000494:pharmacology; D006150:Guanine Nucleotides / Q000378:metabolism*; D006153:Guanosine Diphosphate / Q000494:pharmacology; D006160:Guanosine Triphosphate / Q000494:pharmacology; D066298:In Vitro Techniques; D008274:Magnesium / Q000494:pharmacology*; D008854:Microscopy, Electron; D011485:Protein Binding / Q000187:drug effects; D014404:Tubulin / Q000378:metabolism*
chemical_list: D002021:Buffers; D005971:Glutamates; D006150:Guanine Nucleotides; D014404:Tubulin; D006153:Guanosine Diphosphate; D006160:Guanosine Triphosphate; D004492:Edetic Acid; D008274:Magnesium