A mutually induced conformational fit underlies Ca2+-directed interactions between calmodulin and the proximal C terminus of KCNQ4 K+ channels

Abstract

Edited by Roger J. Colbran Calmodulin (CaM) conveys intracellular Ca 2؉ signals to KCNQ (Kv7, "M-type") K ؉ channels and many other ion channels. Whether this "calmodulation" involves a dramatic structural rearrangement or only slight perturbations of the CaM/ KCNQ complex is as yet unclear. A consensus structural model of conformational shifts occurring between low nanomolar and physiologically high intracellular [Ca 2؉ ] is still under debate. Here, we used various techniques of biophysical chemical analyses to investigate the interactions between CaM and synthetic peptides corresponding to the A and B domains of the KCNQ4subtype. We found that in the absence of CaM, the peptides are disordered, whereas Ca 2؉ /CaM imposed helical structure on both KCNQ A and B domains. Isothermal titration calorimetry revealed that Ca 2؉ /CaM has higher affinity for the B domain than for the A domain of KCNQ2-4 and much higher affinity for the B domain when prebound with the A domain. X-ray crystallography confirmed that these discrete peptides spontaneously form a complex with Ca 2؉ /CaM, similar to previous reports of CaM binding KCNQ-AB domains that are linked together. Microscale thermophoresis and heteronuclear singlequantum coherence NMR spectroscopy indicated the C-lobe of Ca 2؉ -free CaM to interact with the KCNQ4 B domain (K d ϳ10 -20 M), with increasing Ca 2؉ molar ratios shifting the CaM-B domain interactions via only the CaM C-lobe to also include the N-lobe. Our findings suggest that in response to increased Ca 2؉ , CaM undergoes lobe switching that imposes a dramatic mutually induced conformational fit to both the proximal C terminus of KCNQ4 channels and CaM, likely underlying Ca 2؉ -dependent regulation of KCNQ gating.

Atomistic Insights of Calmodulin Gating of Complete Ion Channels

“…Several compounds that enhance SK function have been co-crystallized with the isolated CaMBD of SK2 [50,51]. Importantly, riluzole, the first medication approved for amyotrophic lateral sclerosis is among them [60][61][62]. The docking site is located at the N-lobe.…”

Coarse-Grained Modeling and Molecular Dynamics Simulations of Ca2+-Calmodulin

“…Upon sufficient increase in Ca 2+ concentration, Ca 2+ -free CaM transitions to the Ca 2+ -loaded CaM ( Figure 1 ), exposing the hydrophobic target-binding surfaces of CaM to the solvent ( Figure 2 ) ( Barton et al, 2002 ; Vetter and Leclerc, 2003 ; Park et al, 2008 ; Gromiha and Gromiha, 2010 ; Wu et al, 2012 ; Fernandes and Oliveira-Brett, 2017 ). This transition is accompanied by large conformational changes mutually induced by the conformational changes in the CaM-binding target (CaMBT) peptides ( Wang et al, 2013 ; Liu et al, 2017a ; Archer et al, 2019 ; Bekker et al, 2020 ). The most unique feature of CaM is its reciprocal relation between Ca 2+ binding and target binding ( Meador et al, 1993 ; Weinstein and Mehler, 1994 ; Wall et al, 1997 ; Wriggers et al, 1998 ; Chin and Means, 2000 ; Hoeflich and Ikura, 2002 ; Westerlund and Delemotte, 2018 ), where the conformational changes in the CaM/CaMBT compound further tune CaM’s affinity for the Ca 2+ ions ( Hoffman et al, 2014 ; Zhang et al, 2017 ).…”

“…Although in separate studies, there have been development in AWSEM force fields on structurally flexible proteins ( Wu et al, 2018 ) or on di-valent ions ( Tsai et al, 2016 ), both features, however, have yet to be combined for modeling CaM. Particularly, the coordination chemistry of Ca 2+ ions and the structural flexibility allowing CaM to flex according to its binding targets and surrounding environment is important in modeling the adaptive behavior of CaM ( Wang et al, 2013 ; Liu et al, 2017a ; Archer et al, 2019 ; Bekker et al, 2020 ). By taking into account the coordination chemistry of the Ca 2+ ions as well as the flexible nature of the central linker of CaM, our proposed model for Ca 2+ -CaM elucidates the many-body effects of calcium binding on the geometrical shape of the calcium-binding loop and allows large conformational changes for binding various targets.…”

Pharmacological Manipulation of Kv7 Channels as a New Therapeutic Tool for Multiple Brain Disorders

“…Since PIP 2 binding is required for M-channel opening (Zhang et al, 2003;Li et al, 2005;Suh et al, 2006;Sun and MacKinnon, 2020), its depletion reduces M-current amplitudes in a voltage-independent manner (Shapiro et al, 2000;Nakajo and Kubo, 2005;Choveau et al, 2018). However, in those same neurons, other G q/11 -coupled receptors suppress M current via release of Ca 2+ from IP 3gated stores, loading of Ca 2+ into calmodulin (Gamper and Shapiro, 2003;Winks et al, 2005;Zaika et al, 2007) and changes in configuration of CaM molecules bound to the proximal carboxy terminus of K v 7.1-7.5 subunits (Yus-Najera et al, 2002;Haitin and Attali, 2008) in varying configurations (Haitin and Attali, 2008;Kosenko and Hoshi, 2013;Strulovich et al, 2016;Sun and MacKinnon, 2017;Chang et al, 2018;Archer et al, 2019).…”