Effect of thermostable mutations on the neurotensin receptor 1 (NTSR 1 ) activation state

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Abstract

Neurotensin (NTS) is a 13-amino acid neuropeptide with neuroendocrine and vasoactive functions that is widely expressed in the central nervous system and gastrointestinal tract. NTS is sensed by a multiple cell surface proteins including two G protein-coupling receptors (GPCRs): NTS receptors 1 and 2 (NTSR 1 and NTSR 2 ). Crystal structures of NTSR 1 have successfully elucidated agonist binding within the orthosteric pocket of receptor but have not revealed the full activation state of the receptor. Recent studies have attempted to address this challenge by improving NTSR 1 crystal formation via thermostable mutants; unfortunately, these mutations exhibit functional defects in the G protein coupling of NTSR 1 . Here, we have used molecular dynamics simulations to gain greater insights into how the amino acid substitutions used in these thermostable mutants (E166A, L310A and F358A) impact receptor activation. Our simulations indicate that wild-type NTSR 1 in complex with NTS 8-13 shows more active-like features including a 17.7 Å shift in TM6, reflecting a network of polar and aromatic interactions orchestrating agonist-induced receptor conformational changes. We also provide evidence indicating that F358 is a precursor to the rotamer change observed in W321, and our collective analysis also suggests that mutations E166A and F358A are less impactful to G protein coupling than L310A. Furthermore, we believe that our findings can be used to design future NTSR 1 mutants that do not interfere with agonist-induced conformational changes and downstream G protein coupling and thus produce structures that will allow visualization of the fully activated receptor conformation. Communicated by Ramaswamy H. Sarma.

Original languageEnglish (US)
JournalJournal of Biomolecular Structure and Dynamics
DOIs
StatePublished - Jan 1 2019

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GTP-Binding Proteins
Neurotensin
Mutation
neurotensin (8-13)
Amino Acid Substitution
Molecular Dynamics Simulation
Neuropeptides
Gastrointestinal Tract
Membrane Proteins
Central Nervous System
Amino Acids
neurotensin type 1 receptor

Keywords

  • GPCR
  • molecular dynamics
  • NTSR
  • receptor activation
  • thermostable mutant

ASJC Scopus subject areas

  • Structural Biology
  • Molecular Biology

Cite this

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title = "Effect of thermostable mutations on the neurotensin receptor 1 (NTSR 1 ) activation state",
abstract = "Neurotensin (NTS) is a 13-amino acid neuropeptide with neuroendocrine and vasoactive functions that is widely expressed in the central nervous system and gastrointestinal tract. NTS is sensed by a multiple cell surface proteins including two G protein-coupling receptors (GPCRs): NTS receptors 1 and 2 (NTSR 1 and NTSR 2 ). Crystal structures of NTSR 1 have successfully elucidated agonist binding within the orthosteric pocket of receptor but have not revealed the full activation state of the receptor. Recent studies have attempted to address this challenge by improving NTSR 1 crystal formation via thermostable mutants; unfortunately, these mutations exhibit functional defects in the G protein coupling of NTSR 1 . Here, we have used molecular dynamics simulations to gain greater insights into how the amino acid substitutions used in these thermostable mutants (E166A, L310A and F358A) impact receptor activation. Our simulations indicate that wild-type NTSR 1 in complex with NTS 8-13 shows more active-like features including a 17.7 {\AA} shift in TM6, reflecting a network of polar and aromatic interactions orchestrating agonist-induced receptor conformational changes. We also provide evidence indicating that F358 is a precursor to the rotamer change observed in W321, and our collective analysis also suggests that mutations E166A and F358A are less impactful to G protein coupling than L310A. Furthermore, we believe that our findings can be used to design future NTSR 1 mutants that do not interfere with agonist-induced conformational changes and downstream G protein coupling and thus produce structures that will allow visualization of the fully activated receptor conformation. Communicated by Ramaswamy H. Sarma.",
keywords = "GPCR, molecular dynamics, NTSR, receptor activation, thermostable mutant",
author = "Shanthi Nagarajan and Nabil Alkayed and Sanjiv Kaul and Anthony Barnes",
year = "2019",
month = "1",
day = "1",
doi = "10.1080/07391102.2019.1573705",
language = "English (US)",
journal = "Journal of Biomolecular Structure and Dynamics",
issn = "0739-1102",
publisher = "Adenine Press",

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TY - JOUR

T1 - Effect of thermostable mutations on the neurotensin receptor 1 (NTSR 1 ) activation state

AU - Nagarajan, Shanthi

AU - Alkayed, Nabil

AU - Kaul, Sanjiv

AU - Barnes, Anthony

PY - 2019/1/1

Y1 - 2019/1/1

N2 - Neurotensin (NTS) is a 13-amino acid neuropeptide with neuroendocrine and vasoactive functions that is widely expressed in the central nervous system and gastrointestinal tract. NTS is sensed by a multiple cell surface proteins including two G protein-coupling receptors (GPCRs): NTS receptors 1 and 2 (NTSR 1 and NTSR 2 ). Crystal structures of NTSR 1 have successfully elucidated agonist binding within the orthosteric pocket of receptor but have not revealed the full activation state of the receptor. Recent studies have attempted to address this challenge by improving NTSR 1 crystal formation via thermostable mutants; unfortunately, these mutations exhibit functional defects in the G protein coupling of NTSR 1 . Here, we have used molecular dynamics simulations to gain greater insights into how the amino acid substitutions used in these thermostable mutants (E166A, L310A and F358A) impact receptor activation. Our simulations indicate that wild-type NTSR 1 in complex with NTS 8-13 shows more active-like features including a 17.7 Å shift in TM6, reflecting a network of polar and aromatic interactions orchestrating agonist-induced receptor conformational changes. We also provide evidence indicating that F358 is a precursor to the rotamer change observed in W321, and our collective analysis also suggests that mutations E166A and F358A are less impactful to G protein coupling than L310A. Furthermore, we believe that our findings can be used to design future NTSR 1 mutants that do not interfere with agonist-induced conformational changes and downstream G protein coupling and thus produce structures that will allow visualization of the fully activated receptor conformation. Communicated by Ramaswamy H. Sarma.

AB - Neurotensin (NTS) is a 13-amino acid neuropeptide with neuroendocrine and vasoactive functions that is widely expressed in the central nervous system and gastrointestinal tract. NTS is sensed by a multiple cell surface proteins including two G protein-coupling receptors (GPCRs): NTS receptors 1 and 2 (NTSR 1 and NTSR 2 ). Crystal structures of NTSR 1 have successfully elucidated agonist binding within the orthosteric pocket of receptor but have not revealed the full activation state of the receptor. Recent studies have attempted to address this challenge by improving NTSR 1 crystal formation via thermostable mutants; unfortunately, these mutations exhibit functional defects in the G protein coupling of NTSR 1 . Here, we have used molecular dynamics simulations to gain greater insights into how the amino acid substitutions used in these thermostable mutants (E166A, L310A and F358A) impact receptor activation. Our simulations indicate that wild-type NTSR 1 in complex with NTS 8-13 shows more active-like features including a 17.7 Å shift in TM6, reflecting a network of polar and aromatic interactions orchestrating agonist-induced receptor conformational changes. We also provide evidence indicating that F358 is a precursor to the rotamer change observed in W321, and our collective analysis also suggests that mutations E166A and F358A are less impactful to G protein coupling than L310A. Furthermore, we believe that our findings can be used to design future NTSR 1 mutants that do not interfere with agonist-induced conformational changes and downstream G protein coupling and thus produce structures that will allow visualization of the fully activated receptor conformation. Communicated by Ramaswamy H. Sarma.

KW - GPCR

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KW - receptor activation

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