Aging of tubulin at neutral pH: the destabilizing effect of vinca alkaloids.
The effect of the vinca alkaloid drugs, vincristine, vinblastine, catharanthine, and vindoline, on the aging process of tubulin has been examined. It was found that addition of vincristine or vinblastine accelerated by a factor of 3-3.5 the transformation of tubulin from the 5.8 S alpha-beta-tubulin dimer to paucidisperse polymers, with an average sedimentation coefficient of 9 S, previously observed in the absence of drugs (V. Prakash and S. N. Timasheff, 1982, J. Mol. Biol. 160, 499-515). This transformation of tubulin from 5.8 S to "9 S" followed pseudo-first-order kinetics whether the starting protein was predominantly dimeric (i.e., at low drug concentration) or self-associated into the reversible linear polymers induced by the vinca alkaloid drugs at high drug concentration (G. C. Na and S. N. Timasheff, 1980, Biochemistry 19, 1355-1365; V. Prakash and S. N. Timasheff, 1985, Biochemistry 24, 5004-5010). Identical kinetics were found in a fluorescence examination of the loss by tubulin of its ability to bind colchicine specifically, indicating that the rate determining step is a protein conformational change that induces a major change in the far uv circular dichroism spectrum of tubulin. The found lack of an effect of dithiothreitol on the aging and aggregation processes is consistent with the irreversible aggregation being due to the intermolecular coalescence of nonpolar patches on the protein. The observations that vincristine binds to aged tubulin and that the aging of tubulin is accompanied by quenching of the tryptophan fluorescence similar to that which occurs on the binding of the vinca drugs has led to the proposal that the vinca alkaloids stabilize the aged conformation of the protein by interacting with nonpolar regions that may be related to the aggregation sites.
Prakash
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Graduate Department of Biochemistry, Brandeis University, Waltham, Massachusetts 02254-9110.
Timasheff
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D000818:Animals; D001923:Brain Chemistry; D002417:Cattle; D002942:Circular Dichroism; D011487:Protein Conformation; D013050:Spectrometry, Fluorescence; D013056:Spectrophotometry, Ultraviolet; D014404:Tubulin / Q000737:chemistry* / Q000187:drug effects; D014461:Ultracentrifugation; D014747:Vinblastine / Q000031:analogs & derivatives / Q000494:pharmacology; D014748:Vinca Alkaloids / Q000494:pharmacology*; D014750:Vincristine / Q000494:pharmacology
D014404:Tubulin; D014748:Vinca Alkaloids; C009667:vindoline; D014750:Vincristine; D014747:Vinblastine; C017836:catharanthine