Linkages between the effects of taxol, colchicine, and GTP on tubulin polymerization.

Howard, William D.; Timasheff, Serge N.

doi:10.1016/s0021-9258(19)57307-0

Cited by 58 publications

(30 citation statements)

References 27 publications

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“…The affinity of Mg2+-GTP for the E site, however, is very high (~108 M-1) (Correia et al, 1987). An observation that is inconsistent with the identity of the E-site GTP-bound magnesium and the ion identified in the current study is the observation that, in the presence of taxol, tubulin polymerizes into microtubules whether the E site is occupied by GTP or GDP (Schiff & Horowitz, 1981;Carlier & Pantaloni, 1983), yet magnesium is required (Howard & Timasheff, 1988;Andreu et al, 1992).…”

Section: Mg Activity (M)contrasting

confidence: 69%

Linkages between the dissociation of .alpha..beta. tubulin into subunits and ligand binding: The ground state of tubulin is the GDP conformation

Shearwin

Pérez-Ramírez²,

Sn³

1994

Biochemistry

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The effects of ligands on the dissociation of the alpha beta tubulin dimer into the two subunits were investigated using calf brain tubulin. Sedimentation equilibrium studies showed a number of linkages. In the absence of magnesium in the medium, tubulin-GTP, tubulin-GDP, and tubulin with the exchangeable site unoccupied associate with essentially the same strength (K alpha beta = 1 x 10(7) M-1). This indicates that the ground state of tubulin (i.e., in the absence of magnesium) is not affected by occupancy of the exchangeable nucleotide binding site (E site). The alpha beta association is enhanced by magnesium ions. The association of tubulin with GDP in the E site is linked to the uptake of twice as many magnesium ions as that of tubulin with GTP in the E site. This suggests that magnesium binding is linked to an E-site-related conformational change. Consideration of the linkages between the binding of magnesium ions, E-site occupancy, and tubulin conformation in terms of the model [Howard, W. D., & Timasheff, S. N. (1986) Biochemistry 25, 8292-8300] in which the tubulin alpha beta dimer exists in an equilibrium between two conformations, a microtubule-forming ("straight") state favored by GTP and a double-ring-forming ("curved") state favored by GDP, leads to the conclusion that the ground state of tubulin is the ring-forming or "curved" conformation. Thus, in the absence of magnesium, the tubulin heterodimer exists in the ring-forming conformation, whether the E site is occupied by GTP or GDP.(ABSTRACT TRUNCATED AT 250 WORDS)

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Section: Mg Activity (M)contrasting

confidence: 69%

Linkages between the dissociation of .alpha..beta. tubulin into subunits and ligand binding: The ground state of tubulin is the GDP conformation

Shearwin

Pérez-Ramírez²,

Sn³

1994

Biochemistry

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“…The mechanism of action of these drugs is to bind ␤-tubulin and affect the stabilization of abnormal arrays of MTs in cells leading to an interference of normal tubulin function, including formation of an effective mitotic spindle. The detailed mechanism of taxoid stabilization remains controversial: binding to tubulin could either strengthen the weak lateral interactions between tubulin subunits (Andreu et al, 1992;Howard and Timasheff, 1988), or improve the longitudinal bonds of MTs as proposed by Dye et al (1992). One of the most significant differences between the ␣and ␤-monomers of tubulin is the deletion of eight-loop residues in the ␤-monomer.…”

Section: Effect Of Pacilitaxel Binding On Tubulin Flexibilitymentioning

confidence: 99%

Relating Molecular Flexibility to Function: A Case Study of Tubulin

Keskin

Durell

Bahar

et al. 2002

Biophysical Journal

148

104

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Microtubules (MT), along with a variety of associated motor proteins, are involved in a range of cellular functions including vesicle movement, chromosome segregation, and cell motility. MTs are assemblies of heterodimeric proteins, alpha beta-tubulins, the structure of which has been determined by electron crystallography of zinc-induced, pacilitaxel-stabilized tubulin sheets. These data provide a basis for examining relationships between structural features and protein function. Here, we study the fluctuation dynamics of the tubulin dimer with the aim of elucidating its functional motions relevant to substrate binding, polymerization/depolymerization and MT assembly. A coarse-grained model, harmonically constrained according to the crystal structure, is used to explore the global dynamics of the dimer. Our results identify six regions of collective motion, comprised of structurally close but discontinuous sequence fragments, observed only in the dimeric form, dimerization being a prerequisite for domain identification. Boundaries between regions of collective motions appear to act as linkages, found primarily within secondary-structure elements that lack sequence conservation, but are located at minima in the fluctuation curve, at positions of hydrophobic residues. Residue fluctuations within these domains identify the most mobile regions as loops involved in recognition of the adjacent regions. The least mobile regions are associated with nucleotide binding sites where lethal mutations occur. The functional coupling of motions between and within regions identifies three global motions: torsional and wobbling movements, en bloc, between the alpha- and beta-tubulin monomers, and stretching longitudinally. Further analysis finds the antitumor drug pacilitaxel (TaxotereR) to reduce flexibility in the M loop of the beta-tubulin monomer; an effect that may contribute to tightening lateral interactions between protofilaments assembled into MTs. Our analysis provides insights into relationships between intramolecular tubulin movements of MT organization and function.

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“…Taxol reversibly binds to microtubules in vitro with a maximum stoichiometry of approximately 1 mol of taxol/ mol of tubulin in microtubules and apparently does not bind to free tubulin (Pamess & Horwitz, 1981;. At concentrations that are stoichiometric or near-stoichiometric with respect to the concentrations of tubulin present in solution, taxol strongly stimulates the rate and extent of microtubule polymerization and reduces the concentration of soluble tubulin (Schiff et al, 1979; Kumar, 1981;Howard & Timasheff, 1988). The increased polymer mass is associated with a large taxol-associated linkage free energy contribution (Howard & Timasheff, 1988;.…”

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confidence: 99%

Substoichiometric Binding of Taxol Suppresses Microtubule Dynamics

Derry¹,

Wilson²,

Jordan³

1995

Biochemistry

324

280

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We have measured the effects of taxol (10 nM to 1 microM) on the growing and shortening dynamics at the ends of individual bovine brain microtubules in vitro and have correlated the effects both with the stoichiometry of taxol binding to tubulin in microtubules and with the changes in the microtubule polymer mass. The results indicate that taxol suppresses microtubule dynamic instability differently depending upon the stoichiometry of taxol binding to the microtubules. At the lowest effective concentrations (< or = 100 nM), substoichiometric binding of taxol to tubulin in microtubules (between 0.001 and 0.01 mol of bound taxol/mol of tubulin in microtubules) potently and selectively suppresses the rate and extent of shortening at plus ends in association with some increase (28% to 60%) in the mass of microtubule polymer. At intermediate taxol concentrations (between 100 nM and 1 microM), the binding of additional taxol molecules to the microtubules (between 0.01 and 0.1 mol of taxol bound/mol of tubulin in microtubules) inhibits both growing and shortening events at both microtubule ends with no additional increase in microtubule polymer mass. At high taxol concentrations and high taxol binding stoichiometries (> or = 1 microM taxol and > or = 0.1 mol of taxol bound/mol of tubulin in microtubules), microtubule mass increases sharply and dynamics is almost completely suppressed. The data support the hypothesis that binding of a molecule of taxol to a tubulin subunit in microtubules induces a conformational change in that subunit that strongly reduces its ability to dissociate when the subunit becomes exposed at the microtubule end.

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Linkages between the effects of taxol, colchicine, and GTP on tubulin polymerization.

Cited by 58 publications

References 27 publications

Linkages between the dissociation of .alpha..beta. tubulin into subunits and ligand binding: The ground state of tubulin is the GDP conformation

Linkages between the dissociation of .alpha..beta. tubulin into subunits and ligand binding: The ground state of tubulin is the GDP conformation

Relating Molecular Flexibility to Function: A Case Study of Tubulin

Substoichiometric Binding of Taxol Suppresses Microtubule Dynamics

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