A Dual Involvement of the Amino-terminal Domain of Ezrin in F- and G-actin Binding*
- From the Laboratoire de Dynamique Moléculaire des Interactions Membranaires, CNRS UMR 5539, Université Montpellier II, Bât. 24, CC107, place Eugène Bataillon, 34095 Montpellier Cedex 5, France
Abstract
Human recombinant ezrin, or truncated forms, were coated in microtiter plate and their capacity to bind actin determined. F-actin bound ezrin with a K d of 504 ± 230 nm and a molecular stoichiometry of 10.6 actin per ezrin. Ezrin bound both α- and β/γ-actin essentially as F-form. F-actin binding was totally prevented or drastically reduced when residues 534–586 or 13–30 were deleted, respectively. An actin binding activity was detected in amino-terminal constructs (ezrin 1–310 and 1–333) provided the glutathione S-transferase moiety of the fusion protein was removed. Series of carboxyl-terminal truncations confirmed the presence of this actin-binding site which bound both F- and G-actin. The F- and G-actin-binding sites were differently sensitive to various chemical effectors and distinct specific ezrin antibodies. The internal actin-binding site was mapped between residues 281 and 333. The association of ezrin amino-terminal fragment to full-length ezrin blocked F-actin binding to ezrin. It is proposed that, in full-length ezrin, the F-actin-binding site required the juxtaposition of the distal-most amino- and carboxyl-terminal residues of the ezrin molecule.
Footnotes
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↵* This work was supported by grants from l’Association pour la Recherche sur le Cancer (contract 6844), la Ligue Nationale contre le Cancer, the European Union (BMH4-CT95 0090), and CNRS (Cell Biology project 96033).The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
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↵‡ To whom correspondence should be addressed: UMR 5539, Université de Montpellier II, CC 107, Bât. 24, Place Eugène Bataillon, 34095 Montpellier Cedex 5, France. Tel.: 33-4-67-14-47-28; Fax: 33-4-67-14-47-27 or 33-4-67-14-42-86; E-mail:roy{at}univ-montp2.fr.
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↵1 The abbreviations used are: ERM, ezrin, radixin, moesin; Ez, ezrin; GST, glutathione S-transferase; N- and C-ERMADs, N- and C-ERM association domains; MES, 4-morpholineethanesulfonic acid.
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↵2 Martin, M. (1997) Mol. Biol. Cell, in press.
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↵3 C. Roy, unpublished data.
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↵4 C. Roy, unpublished results.
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- Received March 14, 1997.
- Revision received May 9, 1997.
