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J. Biol. Chem., Vol. 282, Issue 23, 17090-17100, June 8, 2007

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Molecular Characterization of the Ran-binding Zinc Finger Domain of Nup153^*

Meda M. Higa¹, Steven L. Alam², Wesley I. Sundquist, and Katharine S. Ullman³

From the Department of Oncological Sciences, Huntsman Cancer Institute and Department of Biochemistry, University of Utah, Salt Lake City, Utah 84112

The nuclear pore complex is the gateway for selective traffic between the nucleus and cytoplasm. To learn how building blocks of the pore can create specific docking sites for transport receptors and regulatory factors, we have studied a zinc finger module present in multiple copies within the nuclear pores of higher eukaryotes. All four zinc fingers of human Nup153 were found to bind the small GTPase Ran with dissociation constants ranging between 5 and 40 µM. In addition a fragment of Nup153 encompassing the four tandem zinc fingers was found to bind Ran with similar affinity. NMR structural studies revealed that a representative Nup153 zinc finger adopts the same zinc ribbon structure as the previously characterized Npl4 NZF module. Ran binding was mediated by a three-amino acid motif (Leu¹³/Val¹⁴/Asn²⁵) located within the two zinc coordination loops. Nup153 ZnFs bound GDP and GTP forms of Ran with similar affinities, indicating that this interaction is not influenced by a nucleotide-dependent conformational switch. Taken together, these studies elucidate the Ran-binding interface on Nup153 and, more broadly, provide insight into the versatility of this zinc finger binding module.

Received for publication, March 29, 2007

^* This work was supported in part by National Institutes of Health Grant PA CA42014 in support of the DNA Synthesis and Mass Spectrometry Core facilities, National Institutes of Health Grants RO1 GM61275 (to K. S. U.) and RO1 AI45405 (to W. I. S.), and the Leukemia and Lymphoma Society (to K. S. U.). The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

The on-line version of this article (available at http://www.jbc.org) contains supplemental Figs. S1 and S2 and Tables S1–S3.

The atomic coordinates and structure factors (code 2GQE) have been deposited in the Protein Data Bank, Research Collaboratory for Structural Bioinformatics, Rutgers University, New Brunswick, NJ (http://www.rcsb.org/).

¹ Supported by National Institutes of Health Training Grant 5 T32 GM07464.

² Supported in part by the American Cancer Society Grant IRG 77-003-23. To whom correspondence may be addressed: 15 N Medical Dr. East, Rm. 4100, University of Utah, Salt Lake City, UT 84132. Tel.: 801-585-0583; Fax: 801-581-7959; E-mail: alam{at}biochem.utah.edu. ³To whom correspondence may be addressed: 2000 Circle of Hope Dr., University of Utah, Salt Lake City, UT 84112. Tel.: 801-581-7123; Fax: 801-585-0900; E-mail: katharine.ullman{at}hci.utah.edu.

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