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Tubulin domains for the interaction of microtubule associated protein DMAP-85 from Drosophila melanogaster

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Abstract

The interaction of microtubule associated proteins (MAPs) with the microtubule system has been characterized in depth in neuronal cells from various mammalian species. These proteins interact with well-defined domains within the acidic tubulin carboxyl-terminal regulatory region. However, there is little information on the mechanisms of MAPs-tubulin interactions in nonmammalian systems. Recently, a novel tau-like protein designated as DMAP-85 has been identified in Drosophila melanogaster, and the regulation of its interactions with cytoskeletal elements was analyzed throughout different developmental stages of this organism. In this report, the topographic domains involved in the binding of DMAP-85 with tubulin heterodimer were investigated. Affinity chromatography of DMAP-85 in matrixes of taxol-stabilized microtubules showed the reversible interaction of DMAP-85 with domains on the microtubular surface. Co-sedimentation studies using the subtilisin-treated tubulin (S-tubulin) indicated the lack of association of DMAP-85 to this tubulin moiety. Moreover, studies on affinity chromatography of the purified 4 kDa C-terminal tubulin peptide bound to an affinity column, confirmed that DMAP-85 interacts directly with this regulatory domain on tubulin subunits. Further studies on sequencial affinity chromatography using a calmodulin affinity column followed by the microtubule column confirmed the similarities in the interaction behavior of DMAP-85 with that of tau. DMAP-85 associated to both calmodulin and the microtubular polymer. These studies support the idea that the carboxyl-terminal region on tubulin constitutes a common binding domain for most microtubule-interacting proteins.

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Abbreviations

MAPs:

microtubule-associated proteins

C-terminal:

carboxyl-terminal

SDS-PAGE:

polyacrylamide gel electrophoresis in the presence of SDS

DTT:

dithiotreitol

BSA:

bovine serum albumin

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Authors and Affiliations

  1. Laboratory of Cellular and Molecular Biology, Faculty of Sciences, University of Chile and International Center for Cancer and Developmental Biology (ICC), Casilla 653, Santiago, Chile

    Juan P. Henríquez, Verónica Cambiazo & Ricardo B. Maccioni

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  1. Juan P. Henríquez
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  2. Verónica Cambiazo
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  3. Ricardo B. Maccioni
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Henríquez, J.P., Cambiazo, V. & Maccioni, R.B. Tubulin domains for the interaction of microtubule associated protein DMAP-85 from Drosophila melanogaster . Mol Cell Biochem 158, 149–159 (1996). https://doi.org/10.1007/BF00225841

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  • DOI: https://doi.org/10.1007/BF00225841

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