J. Biol. Chem., Vol. 277, Issue 49, 47885-47890, December 6, 2002 Zinc Mediates Assembly of the T1 Domain of the Voltage-gated K Channel 4.2* Alex W. Jahng 1§, Candace Strang ¶, Don Kaiser 1a, Thomas Pollard 1**, Paul Pfaffinger¶, and Senyon Choe 1b From the 1-Structural Biology Laboratory, The Salk Institute, La Jolla, California 92037 and the ¶ Division of Neuroscience, S613, Baylor College of Medicine, 1 Baylor Plaza, Houston, Texas 77030 An intermolecular Zn2+-binding site was identified in the structure of the T1 domain of the Shaw-type potassium channels (aKv3.1). T1 is a BTB/POZ-type domain responsible for the ordered assembly of voltage-gated potassium channels and interactions with other macromolecules. In this structure, a Zn2+ ion was found to be coordinated between each of the four assembly interfaces of the T1 tetramer by three Cys and one His encoded in the sequence motif (HX5CX20CC) of the T1 domain. This sequence motif is conserved among all non-Shaker-type voltage-dependent potassium (Kv) channels, but not in Shaker-type channels. The presence of this conserved Zn2+-binding site is a primary molecular determinant that distinguishes the tetrameric assembly of non-Shaker Kv channel subunits from that of Shaker channels. We report here that tetramerization of the Shal (rKv4.2) T1 in solution requires the presence of Zn2+, and the addition/removal of Zn2+ reversibly switches the protein between a stable tetrameric or monomeric state. We further show that the conversion from tetramers to monomers is profoundly pH-dependent: as the solution pH gets lower, the dissociation rate increases significantly. The unfolding energy of the T1 tetramer as a measure of the conformational stability of the structure is also pH-dependent. Surprisingly, at a lower pH we observe a distinctly altered conformational state of the T1 tetramer trapped during the process of unfolding of the T1 tetramer in the presence of Zn2+. The conformational alteration may be responsible for increased rate of dissociation at lower pH by allowing Zn2+ to be removed more effectively by EDTA. The ability of the T1 domain to adopt stable alternative conformations may be essential to its function as a protein-protein interaction/signaling domain to modulate the ion conduction properties of intact full-length Kv channels. -------------------------------------------------------------------------------- *-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. §-Present address: University of California, Irvine, CA 92697. a-Present address: The Burnham Inst., La Jolla, CA 92037. **-Present address: Dept. of Molecular, Cellular, and Developmental Biology, Yale University, New Haven, CT 06520. b-To whom correspondence should be addressed. Fax: 858-452-3683; E-mail: choe@salk.edu. -------------------------------------------------------------------------------- Copyright © 2002 by The American Society for Biochemistry and Molecular Biology, Inc.