Visualization of arrestin recruitment by a G-protein-coupled receptor

被引：395

作者：

Shukla, Arun K. ^{[1
]}

Westfield, Gerwin H. ^{[2
,3
]}

Xiao, Kunhong ^{[1
]}

Reis, Rosana I. ^{[1
]}

Huang, Li-Yin ^{[1
]}

Tripathi-Shukla, Prachi ^{[1
]}

Qian, Jiang ^{[1
]}

Li, Sheng ^{[4
]}

Blanc, Adi ^{[1
]}

Oleskie, Austin N. ^{[2
,3
]}

Dosey, Anne M. ^{[2
,3
]}

Su, Min ^{[2
,3
]}

Liang, Cui-Rong ^{[5
]}

Gu, Ling-Ling ^{[5
]}

Shan, Jin-Ming ^{[5
]}

Chen, Xin ^{[5
]}

Hanna, Rachel ^{[6
]}

Choi, Minjung ^{[7
]}

Yao, Xiao Jie ^{[1
]}

Klink, Bjoern U. ^{[1
]}

Kahsai, Alem W. ^{[1
]}

Sidhu, Sachdev S. ^{[6
]}

Koide, Shohei ^{[8
]}

Penczek, Pawel A. ^{[9
]}

Kossiakoff, Anthony A. ^{[8
]}

Woods, Virgil L., Jr. ^{[4
]}

Kobilka, Brian K. ^{[10
]}

Skiniotis, Georgios ^{[2
,3
]}

Lefkowitz, Robert J. ^{[1
,7
,11
]}

机构：

[1] Duke Univ, Med Ctr, Dept Med, Durham, NC 27710 USA

[2] Univ Michigan, Sch Med, Life Sci Inst, Ann Arbor, MI 48109 USA

[3] Univ Michigan, Sch Med, Dept Biol Chem, Ann Arbor, MI 48109 USA

[4] Univ Calif San Diego, Dept Chem, La Jolla, CA 92093 USA

[5] Changzhou Univ, Sch Pharmaceut & Life Sci, Changzhou 213164, Jiangsu, Peoples R China

[6] Univ Toronto, Terrence Donnelly Ctr Cellular & Biomol Res, Toronto, ON M5S 3E1, Canada

[7] Duke Univ, Med Ctr, Dept Biochem, Durham, NC 27710 USA

[8] Univ Chicago, Dept Biochem & Mol Biol, Chicago, IL 60637 USA

[9] Univ Texas Houston, Med Sch Houston, Dept Biochem & Mol Biol, Houston, TX 77054 USA

[10] Stanford Univ, Sch Med, Dept Mol & Cellular Physiol, Stanford, CA 94305 USA

[11] Duke Univ, Med Ctr, Howard Hughes Med Inst, Durham, NC 27710 USA

来源：

NATURE | 2014年 / 512卷 / 7513期

基金：

美国国家卫生研究院; 加拿大健康研究院;

关键词：

BETA(2) ADRENERGIC-RECEPTOR; CONFORMATIONAL-CHANGES; ELECTRON-MICROSCOPY; CRYSTAL-STRUCTURE; VISUAL ARRESTIN; ACTIVATION; COMPLEX; BETA-ARRESTIN1; IMAGES; DOMAIN;

D O I：

10.1038/nature13430

中图分类号：

O [数理科学和化学]; P [天文学、地球科学]; Q [生物科学]; N [自然科学总论];

学科分类号：

07 ; 0710 ; 09 ;

摘要：

G-protein-coupled receptors (GPCRs) are critically regulated by beta-arrestins, which not only desensitize G-protein signalling but also initiate a G-protein-independent wave of signalling(1-5). A recent surge of structural data on a number of GPCRs, including the beta(2) adrenergic receptor (beta(2)AR)-G-proteincomplex, has provided novel insights into the structural basis of receptor activation(6-11). However, complementary information has been lacking on the recruitment of beta-arrestins to activated GPCRs, primarily owing to challenges in obtaining stable receptor-beta-arrestin complexes for structural studies. Here we devised a strategy for forming and purifying a functional human beta(2)AR-beta-arrestin-1 complex that allowed us to visualize its architecture by single-particle negative-stain electron microscopy and to characterize the interactions between beta(2)AR and beta-arrestin 1 using hydrogen-deuterium exchange mass spectrometry (HDX-MS) and chemical crosslinking. Electron microscopy two-dimensional averages and three dimensional reconstructions reveal bimodal binding of beta-arrestin 1 to the beta(2)AR, involving two separate sets of interactions, one with the phosphorylated carboxy terminus of the receptor and the other with its seven-transmembrane core. Areas of reduced HDX together with identification of crosslinked residues suggest engagement of the finger loop of beta-arrestin 1 with the seven-transmembrane core of the receptor. In contrast, focal areas of raised HDX levels indicate regions of increased dynamics in both the N and C domains of beta-arrestin 1 when coupled to the beta(2)AR. A molecular model of the beta(2)AR-beta-arrestin signalling complex was made by docking activated beta-arrestin 1 and beta(2)AR crystal structures into the electron microscopymap densities with constraints provided by HDX-MS and crosslinking, allowing us to obtain valuable insights into the overall architecture of a receptor-arrestin complex. The dynamic and structural information presented here provides a framework for better understanding the basis of GPCR regulation by arrestins.

引用

页码：218 / +

页数：17

共 36 条

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