Interaction of reduced glutathione with bovine brain tubulin

Incubation of phosphocellulose-purified tubulin with GSH at 30 degrees C results in an inhibition of colchicine binding activity. GSSG has a protective effect against the GSH-induced loss of colchicine-binding. Incubation of tubulin with GSH at 30 degrees C results in the formation of abnormal tubulin polymers which are insensitive to cold. Such aggregation is insensitive to antimicrotubular drugs. Aggregation is inhibited by GSSG but not by DTT or mercaptoethanol. GSH-induced aggregation is very sensitive to the ionic strength of the assembly medium; both the aggregation and colchicine binding inhibition induced by GSH are inhibited at higher ionic strength. These results indicate a very complex interaction of GSH with tubulin.     

Enzymatic Protein Carboxyl Methylation in Rat Posterior Pituitary: Neurophysins in Rapid-Turnover Pool Determine Methyl Accepting Capacity

In vitro stimulation of intact rat posterior pituitary by either veratridine or K+ depolarization results in the concomitant release of neurophysins and in a decrease (70-80%) in their carboxyl methylation as measured either with L-[methyl-3H]methionine in the intact lobes after stimulation or in their homogenates with [methyl-3H]S-adenosyl-L-methionine and purified protein carboxyl methyltransferase. A similar reduction in neurophysin methylation (60%) was observed when the arrival of newly synthesized neurophysins at the posterior pituitary was blocked by colchicine. Experimental data indicate that the reduction in neurophysin content of the lobes after 12 h of colchicine treatment (less than 7%) or after in vitro stimulation (about 10%) cannot account for the marked reduction in neurophysin methylation. The results suggest that the granule pool characterized by rapid turnover of neurophysins probably represents the major source of methyl acceptor proteins in the lobe. In spite of the marked reduction in neurophysin methyl accepting capacity observed after stimulation, there was no parallel increase in methyl accepting capacity of the released neurophysins. We propose that a neurophysin subfraction that might be associated with the membrane of releasable granules participates in the methylation reaction in situ.