Colchicine binding to tubulin from brain homogenates in the presence of the 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 A1(III), Co(II), Ni(II), while compounds structurally related to glycero (glucose and sucrose) did not inhibition 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 latterwere 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 madeinsoluble 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 streteched structures, 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

Effect of low body temperature of the microtubles of toad brain and testes studied with a morphometric and a biochemical method

El análisis morfométrico aplicado a fibras preterminales de cerebro y a células de Sertoli de testículo del sapo Bufo arenarum mostraron que los microtúbos desaparecen en un 75% y 92% respectivamente cuando los animales son enfriados a 1-2-C por 90 minuots. Las estimaciones hechas en cuerpos neuronales de cerebro y en otras células del testículo mostraron los mismos resultados. Trabajos previos de los autores (López y Bertini, 1982) mostraron que el grado de polimerización de tubulina (porcentaje de tubulina insoluble) en sapos enfriados, solo decreció en un 40%. Se postula que el elevado porcentaje de tubulina insolubre en animales enfriados es decreció en un 40%. Se postula que el elevado porcentaje de tubulina insoluble en animales enfriados es debido a la existencia de un pool de tubulina no estructurada en microtúbulos, unida a estructuras membranosas en sitios de alta afinidad y/o a oligómeros de tubulina no reconocible como microtúbulos por microscopía electrónica