Modulation of the Interaction between Neurotensin Receptor NTS1 and Gq Protein by Lipid

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• 作者：Sayaka Inagaki¹ ; Rodolfo Ghirlando² ; Jim F. White¹ ; Jelena Gvozdenovic-Jeremic¹ ; John K. Northup³ ; Reinhard Grisshammer¹ ; ^{rkgriss@helix.nih.gov}
• 关键词：[3H]NT ; [3H]neurotensin ; BSA ; bovine serum albumin ; CDCl3 ; deuterated chloroform ; Chaps ; 3-[(3-cholamidopropyl)dimethylammonio]propanesulfonic acid ; DLS ; dynamic light scattering ; EDTA ; ethylenediaminetetraacetic acid ; GPCR ; G-protein-coupled receptor ; MB
• 刊名：Journal of Molecular Biology
• 出版年：2012
• 期刊代码：102_00222836
• 类别：imm
• 出版时间：16 March, 2012
• 卷：417
• 期：1-2
• 页码：95-111
• 文件大小：1160 K

摘要

Membrane lipids have been implicated to influence the activity of G-protein-coupled receptors (GPCRs). Almost all of our knowledge on the role of lipids on GPCR and G protein function comes from work on the visual pigment rhodopsin and its G protein transducin, which reside in a highly specialized membrane environment. Thus, insight gained from rhodopsin signaling may not be simply translated to other nonvisual GPCRs. Here, we investigated the effect of lipid head group charges on the signal transduction properties of the class A GPCR neurotensin (NT) receptor 1 (NTS1) under defined experimental conditions, using self-assembled phospholipid nanodiscs prepared with the zwitter-ionic lipid 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC), the negatively charged 1-palmitoyl-2-oleoyl-sn-glycero-3-phospho-(1鈥?rac-glycerol) (POPG), or a POPC/POPG mixture. A combination of dynamic light scattering and sedimentation velocity showed that NTS1 was monomeric in POPC-, POPC/POPG-, and POPG-nanodiscs. Binding of the agonist NT to NTS1 occurred with similar affinities and was essentially unaffected by the phospholipid composition. In contrast, Gq protein coupling to NTS1 in various lipid nanodiscs was significantly different, and the apparent affinity of G伪q and G尾₁纬₁ to activated NTS1 increased with increasing POPG content. NTS1-catalyzed GDP/GTP纬S nucleotide exchange at G伪q in the presence of G尾₁纬₁ and NT was crucially affected by the lipid type, with exchange rates higher by 1 or 2 orders of magnitude in POPC/POPG- and POPG-nanodiscs, respectively, compared to POPC-nanodiscs. Our data demonstrate that negatively charged lipids in the immediate vicinity of a nonvisual GPCR modulate the G-protein-coupling step.