Colchicine binding to tubulin from brain homogenates in the presence of sugars, glycols and metal ions. Effect of nickel ions on the tubulin solubility.
The fact that glycerol preserves microtubules from depolymerizing in vitro, and that some ions such as Ca(II) and Mg(II), regulate the assembly-disassembly process of these structures, induced us to study the effect of several sugars, glycols and metal ions on solubility and colchicine affinity of tubulin in rat brain homogenates, and of purified microtubular protein. Inhibition of colchicine binding was significant with glycerol, polyethylene glycol 1000 (PEG-2) and the ions Al(III), Co(II), Ni(II), while compounds structurally related to glycerol (glucose and sucrose) did not inhibit it. Mannitol, instead, increased the activity a 47% over control. Apparently the presence of some compounds in brain homogenates [PEG-2 (1000) and Ni(II)] favored tubulin sedimentation when these latter were centrifuged at 100,000 x g for 150 min at 20 degrees C, but the form in which tubulin becomes aggregated in the pellet is unknown. Nickel ion made insoluble microtubular protein of homogenates and the purified one by more than 90% without causing significant inhibition of the colchicine binding. The sediment containing nickel-treated two cycles purified microtubular protein observed with the electron microscope did not present microtubules, but it revealed the presence of irregular, wavy and stretched structures bearing highly dense dotted material. The sediments became soluble in phosphate-glutamate buffer (pH 6.8) and, when incubated in polymerizing conditions, gave rise to microtubules undistinguishable from those prepared with untreated purified protein.
Beron
W
W
Instituto de Histología y Embriología (IHEM), Facultad de Ciencias Médicas, Universidad Nacional de Cuyo, Mendoza, Argentina.
Bertini
F
F
D000535:Aluminum / Q000494:pharmacology; D000818:Animals; D001923:Brain Chemistry; D002241:Carbohydrates / Q000494:pharmacology*; D002412:Cations / Q000494:pharmacology*; D011232:Chemical Precipitation; D003035:Cobalt / Q000494:pharmacology; D003078:Colchicine / Q000378:metabolism*; D005260:Female; D005404:Fixatives / Q000494:pharmacology; D006018:Glycols / Q000494:pharmacology*; D008870:Microtubules / Q000187:drug effects; D009419:Nerve Tissue Proteins / Q000378:metabolism; D009532:Nickel / Q000494:pharmacology*; D011108:Polymers; D011485:Protein Binding / Q000187:drug effects; D051381:Rats; D012995:Solubility; D014404:Tubulin / Q000378:metabolism*
D002241:Carbohydrates; D002412:Cations; D005404:Fixatives; D006018:Glycols; D009419:Nerve Tissue Proteins; D011108:Polymers; D014404:Tubulin; D003035:Cobalt; D009532:Nickel; D000535:Aluminum; D003078:Colchicine