Structural Flexibility of CaV1.2 and CaV2.2 I-II Proximal Linker Fragments in Solution

Abstract

Voltage-dependent calcium channels (CaV) enable the inward flow of calcium currents for a wide range of cells. CaV1 and CaV2 subtype α1 subunits form the conducting pore using four repeated membrane domains connected by intracellular linkers. The domain I-II linker connects to the membrane gate (IS6), forming an α-helix, and is bound to the CaVβ subunit. Previous studies indicated that this region may or may not form a continuous helix depending on the CaV subtype, thereby modulating channel activation and inactivation properties. Here, we used small-angle x-ray scattering and ensemble modeling analysis to investigate the solution structure of these linkers, extending from the membrane domain and including the CaVβ-binding site, called the proximal linker (PL). The results demonstrate that the CaV1.2 PL is more flexible than the CaV2.2 PL, the flexibility is intrinsic and not dependent on CaVβ binding, and the flexibility can be most easily explained by the presence of conserved glycines. Our analysis also provides a robust example of investigating protein domains in which flexibility plays an essential role.

Order and disorder in intermediate filament proteins

“…S1). Such high content would promote structural flexibility, as was previously shown in other systems[156]. This trend may have a role in enabling the formation of relatively uniform filaments, despite the varying head lengths.…”

A CACNA1C variant associated with cardiac arrhythmias provides mechanistic insights in the calmodulation of L-type Ca2+ channels

“…Increasing flexibility by inserting glycine residues ( 29 , 30 ) decreases channel function. In contrast, decreasing flexibility of this region by removing glycine residues promoted channel function ( 28 , 31 , 79 , 80 ). The presence of a glycine residue proximal to the α-interacting domain in Ca V 1.2 WT could thus explain the requirement of a stronger depolarization in Ca V 1.2 WT versus G449R channels.…”

“…The relative rigidity α-helix could promote a strong van der Waals interaction between the guanylate domain of Ca V β and hydrophobic residues of Ca V 1.2 ( 29 , 83 , 84 ). In the native protein, this α-helix breaks at the glycine located at position 449 ( 79 ). Crystallographic and circular dichroism spectroscopic studies demonstrated that the arginine substitution prolongs the α-helix ( 31 ).…”

Intrinsically Disordered Proteins at the Nano-scale

“…A simple SAXS analysis immediately provides R g and the pair distance distribution [63,64]. Additionally, traditional Kratky plots (q 2 I(q) vs. q), directly obtained from the SAXS profiles of IDPs, provide a qualitative picture of the presence of globular or unfolded conformations [65,66]. Here, I is the scattering intensity, and q is the scattered wave-vector proportional to the scattering angle.…”