Characterization of G protein coupling mediated by the conserved D134^3.49 of DRY motif, M241^6.34, and F251^6.44 residues on human CXCR1

Abstract

HighlightsMutations in the DRY motif of CXCR1 abolish ligand binding and receptor activation.Point mutations between TM6 and i3 loop result in constitutive activity of CXCR1.Constitutive activity of mutant CXCR1 occurs via Gα15 signaling activation.The highly conserved DRY motifs have distinct roles in CXCR1 and CXCR2.

Structural Analysis of Chemokine Receptor–Ligand Interactions

“…The binding mode of Nb7 is similar to other reported nanobody-bound GPCR structures. 85 , 86 The conserved R129 3.50 of the DRY motif in US28 ( Figure 2 ) interacts with Nb7 via van der Waals interactions and water-mediated H-bond interactions with the side chain and backbone of Ile, 101 respectively ( Figure 6 b). Similarly, R131 3.50 of the β 2 -adrenoceptor (ADRB 2 ) interacts with Tyr 391 of the alpha subunit of the Gs-protein (PDB 3SN6) 87 as well as R102 3.50 of the A 2A adenosine receptor does with Tyr 391 of the engineered mini G-protein in a recently published structure (PDB 5G53); 88 R135 3.50 of rhodopsin also interacts with Gly 2076 and Leu 2077 of a visual arrestin (PDB 4ZWJ).…”

Lactobacillus plantarum CQPC06 Activity Prevents Dextran Sulfate Sodium‐Induced Colitis by Regulating the IL‐8 Pathway

“…When Table 1-Sequences of primers used for reverse transcriptionpolymerase chain reactions in this study. nNOS Forward: 5 -TCCCAGTAACGGACCTCAG-3 Reverse: 5 -TGCTCAACACAGGTTCTATCTCT-3 eNOS Forward: 5 -TGTGACCCTCACCGCTACAA-3 Reverse: 5 -GCACAATCCAGGCCCAATC-3 iNOS Forward: 5 -GGAGTGACGGCAAACATGACT-3 Reverse: 5 -TCGATGCACAACTGGGTGAAC-3 NF-κB F o r w a r d : 5 -GGGGCCTGCAAAGGTTATC-3 Reverse: 5 -TGCTGTTACGGTGCATACCC-3 IκB-α Forward: 5 -TGAAGGACGAGGAGTACGAGC-3 Reverse: 5 -TGCAGGAACGAGTCTCCGT-3 CXCR1 Forward: 5 -TAGAAGGTGTTGAGCGGGAAG-3 Reverse: 5 -TGAGACGAGAAGGAGCATTGG-3 CXCR2 Forward: 5 -CTGCCTTAACCCCATCATCT-3 Reverse: 5 -GCCATGCTGAAAGACAAGAA-3 GAPDH Forward: 5 -AGGTCGGTGTGAACGGATTTG-3 Reverse: 5 -GGGGTCGTTGATGGCAACA-3 IL-8 combines with CXCR1/2, CXCR1/2 transduces signals via G-protein coupled receptors (Han, Feng, Chen, Gerard, & Boisvert, 2015). CXCR1 and CXCR2 are primarily expressed on the surfaces of T cells, monocytes, monocytoid B cells, melanoma cells, synovial fibroblasts, neutrophils, and myeloid progenitor cells (Williams et al, 2000).…”

“…Interleukin 8 (IL‐8) was the first CXC chemokine discovered, and is an important ligand for C‐X‐C motif chemokine receptor 1 (CXCR1) and CXCR2. When IL‐8 combines with CXCR1/2, CXCR1/2 transduces signals via G‐protein coupled receptors (Han, Feng, Chen, Gerard, & Boisvert, ). CXCR1 and CXCR2 are primarily expressed on the surfaces of T cells, monocytes, monocytoid B cells, melanoma cells, synovial fibroblasts, neutrophils, and myeloid progenitor cells (Williams et al., ).…”

The DRF motif of CXCR6 as chemokine receptor adaptation to adhesion

“…The highly conserved aspartate-arginine-tyrosine (DRY) motif, located at the cytoplasmic side of transmembrane helix 3 (TM3) of most class A GPCRs, is a key motif for stabilizing the active state of the receptor and to activate G proteins, thereby regulating receptor activity [ 19 – 21 ]. Specifically, the negatively charged D 3.49 (the number in superscript represents the position of the residue in the sequence according to the generic GPCRdb numbering [ 22 ]) forms a salt bridge with the positively charged R 3.50 which keeps this arginine warped in an inactive conformation.…”

“…Specifically, the negatively charged D 3.49 (the number in superscript represents the position of the residue in the sequence according to the generic GPCRdb numbering [ 22 ]) forms a salt bridge with the positively charged R 3.50 which keeps this arginine warped in an inactive conformation. Therefore, D 3.49 has been shown to be involved in regulating the activity of many GPCRs including the chemokine receptors CXCR1, CXCR2, and chemokine (C-C motif) receptor 5 CCR5 [ 19 – 21 ]. Upon receptor activation, R 3.50 is released from its interaction with D 3.49 and extends to interact with Y 5.58 and Y 7.53 , stabilizing the active state and building the binding pocket for the Gα-subunit of the G protein [ 23 , 24 ].…”