Dodd, R; Peracchia, C; Stolady, D; Török, K
(2008)
Calmodulin association with connexin32-derived peptides suggests trans-domain interaction in chemical gating of gap junction channels.
JOURNAL OF BIOLOGICAL CHEMISTRY, 283 (40).
26911 - 26920.
ISSN 0021-9258
https://doi.org/10.1074/jbc.M801434200
SGUL Authors: Torok, Katalin
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Abstract
Calmodulin plays a key role in the chemical gating of gap junction channels. Two calmodulin-binding regions have previously been identified in connexin32 gap junction protein, one in the N-terminal and another in the C-terminal cytoplasmic tail of the molecule. The aim of this study was to better understand how calmodulin interacts with the connexin32-binding domains. Lobe-specific interactions of calmodulin with connexin32 peptides were studied by stopped flow kinetics, using Ca2+ binding-deficient mutants. Peptides corresponding to the N-terminal tail (residues 1–22) of connexin32 engaged both the N- and C-terminal lobes (N- and C-lobes) of calmodulin, binding with higher affinity to the C-lobe of calmodulin (Ca2+ dissociation rate constants k3,4, 1.7 ± 0.5 s–1) than to the N-lobe (k1,2, 10.8 ± 1.3 s–1). In contrast, peptides representing the C-terminal tail domain (residues 208–227) of connexin32 bound either the C- or the N-lobe but only one calmodulin lobe at a time (k3,4, 2.6 ± 0.1 s–1 or k1, 13.8 ± 0.5 s–1 and k2, 1000 s–1). The calmodulin-binding domains of the N- and C-terminal tails of connexin32 were best defined as residues 1–21 and 216–227, respectively. Our data, showing separate functions of the N- and C-lobes of calmodulin in the interactions with connexin32, suggest trans-domain or trans-subunit bridging by calmodulin as a possible mechanism of gap junction gating.
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