Dataset Information

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 ?1 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 ?-helix, and is bound to the CaV? 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 CaV?-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 CaV? 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.

SUBMITTER: Almagor L

PROVIDER: S-EPMC3672870 | biostudies-other | 2013 Jun

REPOSITORIES: biostudies-other

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Structural flexibility of CaV1.2 and CaV2.2 I-II proximal linker fragments in solution.

Almagor Lior L Avinery Ram R Hirsch Joel A JA Beck Roy R

Biophysical journal 20130601 11

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 act ...[more]

PMID: 23746511

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Project description:Calmodulin (CaM) is a highly-expressed Ca2+ binding protein known to bind hundreds of protein targets. Its binding selectivity to many of these targets is partially attributed to the protein's flexible alpha helical linker that connects its N- and C-domains. It is not well established how its linker mediates CaM's binding to regulatory targets yet. Insights into this would be invaluable to understanding its regulation of diverse cellular signaling pathways. Therefore, we utilized Martini coarse-grained (CG) molecular dynamics simulations to probe CaM/target assembly for a model system: CaM binding to the calcineurin (CaN) regulatory domain. The simulations were conducted assuming a 'wild-type' calmodulin with normal flexibility of its linker, as well as a labile, highly-flexible linker variant to emulate structural changes that could be induced, for instance, by post-translational modifications. For the wild-type model, 98% of the 600 simulations across three ionic strengths adopted a bound complex within 2 μs of simulation time; of these, 1.7% sampled the fully-bound state observed in the experimentally-determined crystallographic structure. By calculating the mean-first-passage-time for these simulations, we estimated the association rate to be ka= 8.7 × 108 M-1 s-1, which is similar to the diffusion-limited, experimentally-determined rate of 2.2 × 108 M-1 s-1. Furthermore, our simulations recapitulated its well-known inverse relationship between the association rate and the solution ionic strength. In contrast, although over 97% of the labile linker simulations formed tightly-bound complexes, only 0.3% achieved the fully-bound configuration. This effect appears to stem from a difference in the ensembles of extended and collapsed states which are controlled by the linker flexibility. Therefore, our simulations suggest that variations in the CaM linker's propensity for alpha helical secondary structure can modulate the kinetics of target binding.

Dataset Information

Structural flexibility of CaV1.2 and CaV2.2 I-II proximal linker fragments in solution.

Publications

Structural flexibility of CaV1.2 and CaV2.2 I-II proximal linker fragments in solution.

Similar Datasets

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