Chemical shift assignments of the connexin45 carboxyl terminal domain: monomer and dimer conformations
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ARTICLE
Chemical shift assignments of the connexin45 carboxyl terminal domain: monomer and dimer conformations Jennifer L. Kopanic • Paul L. Sorgen
Received: 11 July 2012 / Accepted: 5 October 2012 Ó Springer Science+Business Media Dordrecht 2012
Abstract Connexin45 (Cx45) is a gap junction protein involved in cell-to-cell communication in the heart and other tissues. Here we report the 1H, 15N, and 13C resonance assignments for the monomer and dimer conformations of the Cx45 carboxyl terminal (Cx45CT) domain and provide evidence of dimerization using diffusion ordered spectroscopy. The predicted secondary structure of the Cx45CT domain based on the chemical shifts identified one region of a-helical structure, which corresponds to the residues that broadened beyond detection in the dimer confirmation. Previous biophysical studies from our laboratory characterizing the CT domain from the other major cardiac connexins, Cx40 and Cx43, suggest that the amount of a-helical content may translate into the ability of a protein to dimerize. Even though the CT domain is thought to be the main regulatory domain of most connexins, the physiological role of CT dimerization is currently unknown. Therefore, these assignments will be useful for determining the intermolecular interactions that mediate Cx45CT dimerization, information that will be used to characterize dimerization in functional channels, as well as characterizing the binding sites for molecular partners involved in Cx45 regulation. Keywords Cx45 Gap junction Dimerization Carboxyl terminus Intrinsically disordered protein Biological context Gap junctions are integral membrane proteins that enable the direct cytoplasmic exchange of ions and low molecular J. L. Kopanic P. L. Sorgen (&) Department of Biochemistry and Molecular Biology, University of Nebraska Medical Center, Omaha, NE 68198, USA e-mail: [email protected]
weight metabolites (\1 kDa) between adjacent cells. They provide a pathway for the propagation and/or amplification of signal transduction cascades triggered by cytokines, growth factors, and other cell signaling molecules involved in growth, regulation, and development. For example, gap junction intercellular communication is a principal determinant of myocardial conduction, and altered expression has been implicated in arrhythmogenesis (van Veen et al. 2006). Gap junctions are formed by the apposition of two connexons from adjacent cells, where each connexon is formed of six connexin proteins. Connexins are tetraspan transmembrane domain proteins with intracellular amino- and carboxyltermini. There are 21 different connexin genes in the human genome with differential spatial-temporal expression throughout the body. In the heart, connexin45 (Cx45), connexin43 (Cx43), and connexin40 (Cx40) are found in distinctive combinations and relative quantities in different, functionally specialized subsets of cardiomyocytes. Though there is significant sequence homology among all connexins, the major divergence in primary structures occurs in the cyto
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