Structure and mechanism of calmodulin binding to a signaling sphingolipid reveal new aspects of lipid‐protein interactions

Abstract

Lipid-protein interactions are rarely characterized at a structural molecular level due to technical difficulties; however, the biological significance of understanding the mechanism of these interactions is outstanding. In this report, we provide mechanistic insight into the inhibitory complex formation of the lipid mediator sphingosylphosphorylcholine with calmodulin, the most central and ubiquitous regulator protein in calcium signaling. We applied crystallographic, thermodynamic, kinetic, and spectroscopic approaches using purified bovine calmodulin and bovine cerebral microsomal fraction to arrive at our conclusions. Here we present 1) a 1.6-Å resolution crystal structure of their complex, in which the sphingolipid occupies the conventional hydrophobic binding site on calmodulin; 2) a peculiar stoichiometry-dependent binding process: at low or high protein-to-lipid ratio calmodulin binds lipid micelles or a few lipid molecules in a compact globular conformation, respectively, and 3) evidence that the sphingolipid displaces calmodulin from its targets on cerebral microsomes. We have ascertained the specificity of the interaction using structurally related lipids as controls. Our observations reveal the structural basis of selective calmodulin inhibition by the sphingolipid. On the basis of the crystallographic and biophysical characterization of the calmodulin-sphingosylphosphorylcholine interaction, we propose a novel lipid-protein binding model, which might be applicable to other interactions as well.

Inter‐domain electron transfer in cellobiose dehydrogenase: modulation by pH and divalent cations

“…The pH profiles of ferricyanide and cyt c therefore give a clear indication that divalent cations support adoption of a closed conformation by the DH and the CYT when the domain interface becomes negatively charged at neutral/alkaline pH. This effect is not caused by the presence of distinct binding sites for divalent cations, as these were not found in our docking models of the DH and the CYT, and the 30–60 m m concentration of dications required to achieve the optimal effect is magnitudes higher than the reported affinities for Ca 2+ specific EF‐hand binding sites, for example (2.1 μ m or 0.49 μ m ). The high concentration required suggests unspecific shielding of close opposing charges at both sides of the domain interface.…”

Applications of isothermal titration calorimetry in pure and applied research—survey of the literature from 2010

“…Protein/peptide interactions with nucleic acids Lipid/membrane interactions Polysaccharide interactions …”

“…(vii) Lipid/membrane interactions. [325][326][327][328][329][330][331][332][333][334][335][336][337][338][339][340][341] (viii) Polysaccharide interactions. (ix) Protein/peptide interactions with polymers and nanoparticles.…”

A Non-Canonical Calmodulin Target Motif Comprising a Polybasic Region and Lipidated Terminal Residue Regulates Localization

“…Interestingly, a structure has been reported of Ca 2+ -CaM bound to a sphingolipid, with no peptidyl moiety attached [59]. CaM adopted a collapsed conformation with open hydrophobic pockets forming a hydrophobic channel that accommodated four molecules of sphingosylphophorylcholine, with its alkyl chains arranged in a parallel orientation.…”