Cholesterol Is a Dose-Dependent Positive Allosteric Modulator of CCR3 Ligand Affinity and G Protein Coupling

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dc.creatorvan Aalst, Evan (TTU)
dc.creatorWylie, Benjamin J. (TTU)
dc.date.accessioned2022-11-23T15:54:59Z
dc.date.available2022-11-23T15:54:59Z
dc.date.issued2021
dc.description© 2021 van Aalst and Wylie. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.en_US
dc.description.abstractCholesterol as an allosteric modulator of G protein-coupled receptor (GPCR) function is well documented. This quintessential mammalian lipid facilitates receptor–ligand interactions and multimerization states. Functionally, this introduces a complicated mechanism for the homeostatic modulation of GPCR signaling. Chemokine receptors are Class A GPCRs responsible for immune cell trafficking through the binding of endogenous peptide ligands. CCR3 is a CC motif chemokine receptor expressed by eosinophils and basophils. It traffics these cells by transducing the signal stimulated by the CC motif chemokine primary messengers 11, 24, and 26. These behaviors are close to the human immunoresponse. Thus, CCR3 is implicated in cancer metastasis and inflammatory conditions. However, there is a paucity of experimental evidence linking the functional states of CCR3 to the molecular mechanisms of cholesterol–receptor cooperativity. In this vein, we present a means to combine codon harmonization and a maltose-binding protein fusion tag to produce CCR3 from E. coli. This technique yields ∼2.6 mg of functional GPCR per liter of minimal media. We leveraged this protein production capability to investigate the effects of cholesterol on CCR3 function in vitro. We found that affinity for the endogenous ligand CCL11 increases in a dose-dependent manner with cholesterol concentration in both styrene:maleic acid lipid particles (SMALPs) and proteoliposomes. This heightened receptor activation directly translates to increased signal transduction as measured by the GTPase activity of the bound G-protein α inhibitory subunit 3 (Gαi3). This work represents a critical step forward in understanding the role of cholesterol-GPCR allostery in regulation of signal transduction.en_US
dc.identifier.citationvan Aalst E and Wylie BJ (2021) Cholesterol Is a Dose-Dependent Positive Allosteric Modulator of CCR3 Ligand Affinity and G Protein Coupling. Front. Mol. Biosci. 8:724603. doi: 10.3389/fmolb.2021.724603en_US
dc.identifier.urihttps://doi.org/10.3389/fmolb.2021.724603
dc.identifier.urihttps://hdl.handle.net/2346/90351
dc.language.isoengen_US
dc.subjectGPCRen_US
dc.subjectCCR3en_US
dc.subjectChemokine Receptoren_US
dc.subjectCCL11en_US
dc.subjectCholesterolen_US
dc.subjectG Proteinen_US
dc.subjectLipid Allosteryen_US
dc.subjectCodon Harmonizationen_US
dc.titleCholesterol Is a Dose-Dependent Positive Allosteric Modulator of CCR3 Ligand Affinity and G Protein Couplingen_US
dc.typeArticleen_US

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