Direct evidence for GTP and GDP-P sub i intermediates in microtubule assembly

Journal Article · Tue Sep 25 04:00:00 EDT 1990 · Biochemistry; (USA)

DOI:https://doi.org/10.1021/bi00490a007· OSTI ID:5920733

Melki, R; Carlier, M F; Pantaloni, D ^[1]

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1. Laboratoire d'Enzymologie dy CNRS, Gif-sur-Yvette (France)

Identification of the kinetic intermediates in GTP hydrolysis on microtubules and characterization of their assembly properties is essential in understanding microtubule dynamics. By using an improved glass filter assay that selectively traps microtubules with a dead time of 2 s and monitoring taxol-induced rapid assembly of microtubules from ({gamma}-{sup 32}P,{sup 3}H)GTP-tubulin 1:1 complex, direct evidence has been obtained for GTP- and GDP-P{sub i}-microtubule transient states in the early stages of the polymerization process. A simple kinetic analysis of GTP hydrolysis on microtubules within two sequential pseudo-first-order processes led to apparent first-order rate constants of 0.065 s{sup {minus}1} for the cleavage of the {gamma}-phosphate and 0.02 s{sup {minus}1} for the liberation of P{sub i}, assuming a simple random model. Apparent rate constants for GTP hydrolysis and P{sub i} release were independent of the composition of the buffer used to polymerize tubulin. The significance of these values with respect to those derived from previous studies from this and other laboratories and the possibility of a vectorial model for GTP hydrolysis are discussed.

• OSTI ID: 5920733
• Journal Information: Biochemistry; (USA), Journal Name: Biochemistry; (USA) Vol. 29:38; ISSN 0006-2960; ISSN BICHA
• Country of Publication: United States
• Language: English

Related Subjects

550601* -- Medicine-- Unsealed Radionuclides in Diagnostics
62 RADIOLOGY AND NUCLEAR MEDICINE
ADSORPTION
AMINES
AROMATICS
AZAARENES
BIOCHEMICAL REACTION KINETICS
CELL CONSTITUENTS
CHEMICAL REACTIONS
DECOMPOSITION
ELECTRON MICROSCOPY
GUANINE
HETEROCYCLIC COMPOUNDS
HYDROGEN COMPOUNDS
HYDROLYSIS
HYDROXY COMPOUNDS
KINETICS
LYSIS
MICROSCOPY
MICROTUBULES
NUCLEOSIDES
NUCLEOTIDES
ORGANIC COMPOUNDS
ORGANIC NITROGEN COMPOUNDS
POLYMERIZATION
PURINES
REACTION KINETICS
RIBOSIDES
SOLVOLYSIS
SORPTION
TRANSMISSION ELECTRON MICROSCOPY
TRITIUM COMPOUNDS