Promotion of Tubulin Assembly by Poorly Soluble Taxol Analogs

Overview

Journal Anal Biochem

Publisher Elsevier

Specialty Biochemistry

Date 2006 Nov 14

PMID 17097592

Citations 8

Authors

Shubhada Sharma

Thota Ganesh

David G I Kingston

Susan Bane

Affiliations

Soon will be listed here.

Abstract

Promotion or inhibition of tubulin assembly into microtubules is the standard in vitro assay for evaluating potential antimicrotubule agents. Many agents to be tested are poorly soluble in aqueous solution and require a cosolvent such as dimethyl sulfoxide (DMSO). However, DMSO itself can promote tubulin assembly, and its inclusion in assays for compounds that induce tubulin assembly complicates interpretation of the results. Substituting GDP for GTP in the exchangeable nucleotide binding site of tubulin produces a less active form of the protein, tubulin-GDP. Here it is shown that tubulin-GDP can be assembled into normal microtubules in DMSO concentrations up to 15% (v/v), and polymerization assays performed under these conditions can be compared with assays run under more standard conditions. Assays for measuring the effective concentration of a ligand for promotion of tubulin assembly (EC(50)), measuring the concentration for inhibition of tubulin assembly (IC(50)) by a colchicine site ligand, and measuring tubulin critical concentrations in the presence of poorly soluble taxol derivatives are illustrated.

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