Interactions of tubulin with guanylyl-(beta-gamma-methylene)diphosphonate. Formation and assembly of a stoichiometric complex.
Complete replacement of the nucleotide on the exchangeable binding site of purified calf brain tubulin by the non-hydrolyzable GTP-analogue guanylyl-(beta,gamma-methylene)diphosphonate (GMPPCP) has been achieved by treatment of tubulin-GDP with phosphodiesterase-free alkaline phosphatase. GMPPCP binds to tubulin with a low affinity relative to GTP or GDP. Binding of the analogue is linked to magnesium ion concentration and, like the binding of other guanine nucleotides, is promoted by high concentrations of glycerol. The complex of pure tubulin and GMPPCP readily assembles at 37 degrees C into microtubules or curled ribbons of protofilaments, depending on buffer composition. Assemblies are cold-reversible at 0-2 degrees C, and multiple reversible assemblies can be observed during repeated heating/cooling cycles.
Seckler
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Graduate Department of Biochemistry, Brandeis University, Waltham, Massachusetts 02254-9110.
Wu
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GM
Timasheff
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D000818:Animals; D001921:Brain / Q000378:metabolism; D002417:Cattle; D002851:Chromatography, High Pressure Liquid; D005990:Glycerol / Q000494:pharmacology; D006150:Guanine Nucleotides / Q000302:isolation & purification; D006153:Guanosine Diphosphate / Q000378:metabolism; D006160:Guanosine Triphosphate / Q000031:analogs & derivatives* / Q000378:metabolism; D007700:Kinetics; D008854:Microscopy, Electron; D011485:Protein Binding; D014404:Tubulin / Q000302:isolation & purification / Q000378:metabolism* / Q000648:ultrastructure
D006150:Guanine Nucleotides; D014404:Tubulin; D006153:Guanosine Diphosphate; C004805:5'-guanylylmethylenebisphosphonate; D006160:Guanosine Triphosphate; D005990:Glycerol