Gs protein peptidomimetics as allosteric modulators of the β2-adrenergic receptor

Lotte Emilie Boyhus; Mia Danielsen; Nina Smidt Bengtson; Micha Ben Achim Kunze; Xavier Kubiak; Tjerk J. Sminia; Jacob Hartvig Løper; Phuong Thu Tran; Kresten Lindorff-Larsen; Søren G.F. Rasmussen; Jesper Mosolff Mathiesen; Daniel Sejer Pedersen

doi:10.1039/c7ra11713b

G_s protein peptidomimetics as allosteric modulators of the β₂-adrenergic receptor

Lotte Emilie Boyhus, Mia Danielsen, Nina Smidt Bengtson, Micha Ben Achim Kunze, Xavier Kubiak, Tjerk J. Sminia, Jacob Hartvig Løper, Phuong Thu Tran, Kresten Lindorff-Larsen, Søren G.F. Rasmussen, Jesper Mosolff Mathiesen^*, Daniel Sejer Pedersen

^*Corresponding author for this work

3 Citations (Scopus)

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Abstract

A series of G_s protein peptidomimetics were designed and synthesised based on the published X-ray crystal structure of the active state β₂-Adrenergic receptor (β₂AR) in complex with the G_s protein (PDB 3SN6). We hypothesised that such peptidomimetics may function as allosteric modulators that target the intracellular G_s protein binding site of the β₂AR. Peptidomimetics were designed to mimic the 15 residue C-Terminal α-helix of the G_s protein and were pre-organised in a helical conformation by (i, i + 4)-stapling using copper catalysed azide alkyne cycloaddition. Linear and stapled peptidomimetics were analysed by circular dichroism (CD) and characterised in a membrane-based cAMP accumulation assay and in a bimane fluorescence assay on purified β₂AR. Several peptidomimetics inhibited agonist isoproterenol (ISO) induced cAMP formation by lowering the ISO maximal efficacy up to 61%. Moreover, some peptidomimetics were found to significantly decrease the potency of ISO up to 39-fold. In the bimane fluorescence assay none of the tested peptidomimetics could stabilise an active-like conformation of β₂AR. Overall, the obtained pharmacological data suggest that some of the peptidomimetics may be able to compete with the native G_s protein for the intracellular binding site to block ISO-induced cAMP formation, but are unable to stabilise an active-like receptor conformation.

Original language	English
Journal	RSC Advances
Volume	8
Issue number	4
Pages (from-to)	2219-2228
Number of pages	10
ISSN	2046-2069
DOIs	https://doi.org/10.1039/c7ra11713b
Publication status	Published - 2018

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@article{779a451aaab342689abd4cded6482c94,

title = "Gs protein peptidomimetics as allosteric modulators of the β2-adrenergic receptor",

abstract = "A series of Gs protein peptidomimetics were designed and synthesised based on the published X-ray crystal structure of the active state β2-Adrenergic receptor (β2AR) in complex with the Gs protein (PDB 3SN6). We hypothesised that such peptidomimetics may function as allosteric modulators that target the intracellular Gs protein binding site of the β2AR. Peptidomimetics were designed to mimic the 15 residue C-Terminal α-helix of the Gs protein and were pre-organised in a helical conformation by (i, i + 4)-stapling using copper catalysed azide alkyne cycloaddition. Linear and stapled peptidomimetics were analysed by circular dichroism (CD) and characterised in a membrane-based cAMP accumulation assay and in a bimane fluorescence assay on purified β2AR. Several peptidomimetics inhibited agonist isoproterenol (ISO) induced cAMP formation by lowering the ISO maximal efficacy up to 61%. Moreover, some peptidomimetics were found to significantly decrease the potency of ISO up to 39-fold. In the bimane fluorescence assay none of the tested peptidomimetics could stabilise an active-like conformation of β2AR. Overall, the obtained pharmacological data suggest that some of the peptidomimetics may be able to compete with the native Gs protein for the intracellular binding site to block ISO-induced cAMP formation, but are unable to stabilise an active-like receptor conformation.",

author = "Boyhus, {Lotte Emilie} and Mia Danielsen and Bengtson, {Nina Smidt} and Kunze, {Micha Ben Achim} and Xavier Kubiak and Sminia, {Tjerk J.} and L{\o}per, {Jacob Hartvig} and Tran, {Phuong Thu} and Kresten Lindorff-Larsen and Rasmussen, {S{\o}ren G.F.} and Mathiesen, {Jesper Mosolff} and Pedersen, {Daniel Sejer}",

year = "2018",

doi = "10.1039/c7ra11713b",

language = "English",

volume = "8",

pages = "2219--2228",

journal = "RSC Advances",

issn = "2046-2069",

publisher = "RSC Publishing",

number = "4",

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TY - JOUR

T1 - Gs protein peptidomimetics as allosteric modulators of the β2-adrenergic receptor

AU - Boyhus, Lotte Emilie

AU - Danielsen, Mia

AU - Bengtson, Nina Smidt

AU - Kunze, Micha Ben Achim

AU - Kubiak, Xavier

AU - Sminia, Tjerk J.

AU - Løper, Jacob Hartvig

AU - Tran, Phuong Thu

AU - Lindorff-Larsen, Kresten

AU - Rasmussen, Søren G.F.

AU - Mathiesen, Jesper Mosolff

AU - Pedersen, Daniel Sejer

PY - 2018

Y1 - 2018

N2 - A series of Gs protein peptidomimetics were designed and synthesised based on the published X-ray crystal structure of the active state β2-Adrenergic receptor (β2AR) in complex with the Gs protein (PDB 3SN6). We hypothesised that such peptidomimetics may function as allosteric modulators that target the intracellular Gs protein binding site of the β2AR. Peptidomimetics were designed to mimic the 15 residue C-Terminal α-helix of the Gs protein and were pre-organised in a helical conformation by (i, i + 4)-stapling using copper catalysed azide alkyne cycloaddition. Linear and stapled peptidomimetics were analysed by circular dichroism (CD) and characterised in a membrane-based cAMP accumulation assay and in a bimane fluorescence assay on purified β2AR. Several peptidomimetics inhibited agonist isoproterenol (ISO) induced cAMP formation by lowering the ISO maximal efficacy up to 61%. Moreover, some peptidomimetics were found to significantly decrease the potency of ISO up to 39-fold. In the bimane fluorescence assay none of the tested peptidomimetics could stabilise an active-like conformation of β2AR. Overall, the obtained pharmacological data suggest that some of the peptidomimetics may be able to compete with the native Gs protein for the intracellular binding site to block ISO-induced cAMP formation, but are unable to stabilise an active-like receptor conformation.

AB - A series of Gs protein peptidomimetics were designed and synthesised based on the published X-ray crystal structure of the active state β2-Adrenergic receptor (β2AR) in complex with the Gs protein (PDB 3SN6). We hypothesised that such peptidomimetics may function as allosteric modulators that target the intracellular Gs protein binding site of the β2AR. Peptidomimetics were designed to mimic the 15 residue C-Terminal α-helix of the Gs protein and were pre-organised in a helical conformation by (i, i + 4)-stapling using copper catalysed azide alkyne cycloaddition. Linear and stapled peptidomimetics were analysed by circular dichroism (CD) and characterised in a membrane-based cAMP accumulation assay and in a bimane fluorescence assay on purified β2AR. Several peptidomimetics inhibited agonist isoproterenol (ISO) induced cAMP formation by lowering the ISO maximal efficacy up to 61%. Moreover, some peptidomimetics were found to significantly decrease the potency of ISO up to 39-fold. In the bimane fluorescence assay none of the tested peptidomimetics could stabilise an active-like conformation of β2AR. Overall, the obtained pharmacological data suggest that some of the peptidomimetics may be able to compete with the native Gs protein for the intracellular binding site to block ISO-induced cAMP formation, but are unable to stabilise an active-like receptor conformation.

U2 - 10.1039/c7ra11713b

DO - 10.1039/c7ra11713b

M3 - Journal article

AN - SCOPUS:85040919810

SN - 2046-2069

VL - 8

SP - 2219

EP - 2228

JO - RSC Advances

JF - RSC Advances

IS - 4

ER -

Gs protein peptidomimetics as allosteric modulators of the β2-adrenergic receptor

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G_s protein peptidomimetics as allosteric modulators of the β₂-adrenergic receptor