Structural insights into the activation of neurokinin 2 receptor by neurokinin A

Sun, Wenjing; Yuan, Qingning; Zhang, Huanhuan; Yang, Fan; Ling, Shenglong; Luo, Yifan; Lv, Pei; Xu, H. Eric; Tian, Changlin; Yin, Wanchao; Shi, Peng

doi:10.1038/s41421-022-00437-8

Cited by 8 publications

(12 citation statements)

References 11 publications

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“…The peptides were purified by reversed‐phase (RP)‐high‐performance liquid chromatography (HPLC) and characterized by mass‐spectrometry (MS) (Figure S1). The addition of a triglycine motif to the N ‐terminus of these tachykinins was not expected to substantially alter their activity, as the region responsible for receptor activation resides in the C ‐terminus (FXGLM‐NH2), while the N‐terminal region contributes to receptor subtype selectivity (Sun et al, 2022). The activities of NKA and SP 6–11 were compared to their triglycine extended analogues.…”

Section: Resultsmentioning

confidence: 99%

Semi‐synthetic nanobody‐ligand conjugates exhibit tunable signaling properties and enhanced transcriptional outputs at neurokinin receptor‐1

Braga Emidio,

Cheloha

2024

Protein Science

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Antibodies have proven highly valuable for therapeutic development; however, they are typically poor candidates for applications that require activation of G protein‐coupled receptors (GPCRs), the largest collection of targets for clinically approved drugs. Nanobodies (Nbs), the smallest antibody fragments retaining full antigen‐binding capacity, have emerged as promising tools for pharmacologic applications, including GPCR modulation. Past work has shown that conjugation of Nbs with ligands can provide GPCR agonists that exhibit improved activity and selectivity compared to their parent ligands. The neurokinin‐1 receptor (NK1R), a GPCR targeted for the treatment of pain, is activated by peptide agonists such as Substance P (SP) and neurokinin A (NKA), which induce signaling through multiple pathways (Gs, Gq and β‐arrestin). In this study, we investigated whether conjugating NK1R ligands with Nbs that bind to a separate location on the receptor would provide chimeric compounds with distinctive signaling properties. We employed sortase A‐mediated ligation to generate several conjugates consisting of Nbs linked to NK1R ligands. Many of these conjugates exhibited divergent and unexpected signaling properties and transcriptional outputs. For example, some Nb‐NKA conjugates showed enhanced receptor binding capacity, high potency partial agonism, prolonged cAMP production, and an increase in transcriptional output associated with Gs signaling; whereas other conjugates were virtually inactive. Nanobody conjugation caused only minor alterations in ligand‐induced upstream Gq signaling with unexpected enhancements in transcriptional (downstream) responses. Our findings underscore the potential of nanobody conjugation for providing compounds with advantageous properties such as biased agonism, prolonged duration of action, and enhanced transcriptional responses. These compounds hold promise not only for facilitating fundamental research on GPCR signal transduction mechanisms but also for the development of more potent and enduring therapeutics.This article is protected by copyright. All rights reserved.

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Section: Resultsmentioning

confidence: 99%

Semi‐synthetic nanobody‐ligand conjugates exhibit tunable signaling properties and enhanced transcriptional outputs at neurokinin receptor‐1

Braga Emidio,

Cheloha

2024

Protein Science

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“…The addition of a triglycine motif to the N -terminus of these tachykinins was not expected to substantially alter their activity, as the region responsible for receptor activation resides in the C -terminus (FXGLM-NH2), while the N-terminal region contributes to receptor subtype selectivity. 18 The activities of NKA and SP 6-11 were compared to their triglycine extended analogues. This comparison showed modest effects for triglycine extension, except for G3SP 6-11 (EC 50 0.3 nM), which was almost 100 times more potent than SP 6-11 (27.5 nM) in signaling through the G q pathway (Figure S2) .…”

Section: Resultsmentioning

confidence: 99%

Semi-synthetic nanobody-ligand conjugates exhibit tunable signaling properties and enhanced transcriptional outputs at neurokinin receptor-1

Emidio,

Cheloha

2023

Preprint

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Antibodies have proven highly valuable for therapeutic development; however, they are typically poor candidates for applications that require activation of G protein-coupled receptors (GPCRs), the largest collection of targets for clinically approved drugs. Nanobodies (Nbs), the smallest antibody fragments retaining full antigen-binding capacity, have emerged as promising tools for pharmacologic applications, including GPCR modulation. Past work has shown that conjugation of Nbs with ligands can provide GPCR agonists that exhibit improved activity and selectivity compared to their parent ligands. The neurokinin-1 receptor (NK1R), a GPCR targeted for the treatment of pain, is activated by peptide agonists such as Substance P (SP) and neurokinin A (NKA), which induce signaling through multiple pathways (Gs, Gqand β-arrestin). In this study, we investigated whether conjugating NK1R ligands with Nbs that bind to a separate location on the receptor would provide chimeric compounds with distinctive signaling properties. We employed sortase A-mediated ligation to generate several conjugates consisting of Nbs linked to NK1R ligands. Many of these conjugates exhibited divergent and unexpected signaling properties and transcriptional outputs. For example, some Nb-NKA conjugates showed enhanced receptor binding capacity, high potency partial agonism, prolonged cAMP production, and an increase in transcriptional output associated with Gssignaling; whereas other conjugates were virtually inactive. Nanobody conjugation caused only minor alterations in ligand-induced upstream Gqsignaling with unexpected enhancements in transcriptional (downstream) responses. Our findings underscore the potential of nanobody conjugation for providing compounds with advantageous properties such as biased agonism, prolonged duration of action, and enhanced transcriptional responses. These compounds hold promise not only for facilitating fundamental research on GPCR signal transduction mechanisms but also for the development of more potent and enduring therapeutics.

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“…Binding selectivity depends on the C-terminal end and the interactions of amino acids of the rest of the molecule with the receptor. A recent cryo-EM structure determination of NK-2R-Gq protein complex bound to NKA [ 60 ] underlies the importance of several residues that secure the position of NKA within de receptor pocket (see Figure 4 D). Lateral chains of residues Y93 and N90 and N97 in TM2 interact with the backbone carbonyl oxygens of Val7 and Leu9.…”

Section: Tachykinin and Calcitonin Peptide Familiesmentioning

confidence: 99%

“…Additionally, the N-terminus of NKA contacts the ECL2 region of NK-2R (D175 forms a salt bridge with Lys2 carbonyl oxygen), and Phe6 further contacts Met28 in the N-terminal part of NK-2R and I285 in TM7 ( Figure 4 D). When Sun et al [ 60 ] compared NK-2R with NK-1R (PDB ID 7RMG, [ 61 ]), they found very similar structures, except for an outward shift of the TM5 in NK-2R of 2.4 Å. Moreover, they identified significant differences in the arrangement of extracellular loops, specially ECL2.…”

Section: Tachykinin and Calcitonin Peptide Familiesmentioning

confidence: 99%

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Peptidergic Systems and Cancer: Focus on Tachykinin and Calcitonin/Calcitonin Gene-Related Peptide Families

Sánchez

Rodríguez

Coveñas

2023

Cancers

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The roles played by the peptides belonging to the tachykinin (neurokinin A and B) and calcitonin/calcitonin gene-related peptide (adrenomedullin, adrenomedullin 2, amylin, and calcitonin gene-related peptide (CGRP)) peptide families in cancer development are reviewed. The structure and dynamics of the neurokinin (NK)-2, NK-3, and CGRP receptors are studied together with the intracellular signaling pathways in which they are involved. These peptides play an important role in many cancers, such as breast cancer, colorectal cancer, glioma, lung cancer, neuroblastoma, oral squamous cell carcinoma, phaeochromocytoma, leukemia, bladder cancer, endometrial cancer, Ewing sarcoma, gastric cancer, liver cancer, melanoma, osteosarcoma, ovarian cancer, pancreatic cancer, prostate cancer, renal carcinoma, and thyroid cancer. These peptides are involved in tumor cell proliferation, migration, metastasis, angiogenesis, and lymphangiogenesis. Several antitumor therapeutic strategies, including peptide receptor antagonists, are discussed. The main research lines to be developed in the future are mentioned.

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Structural insights into the activation of neurokinin 2 receptor by neurokinin A

Cited by 8 publications

References 11 publications

Semi‐synthetic nanobody‐ligand conjugates exhibit tunable signaling properties and enhanced transcriptional outputs at neurokinin receptor‐1

Semi‐synthetic nanobody‐ligand conjugates exhibit tunable signaling properties and enhanced transcriptional outputs at neurokinin receptor‐1

Semi-synthetic nanobody-ligand conjugates exhibit tunable signaling properties and enhanced transcriptional outputs at neurokinin receptor-1

Peptidergic Systems and Cancer: Focus on Tachykinin and Calcitonin/Calcitonin Gene-Related Peptide Families

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