Rational Design of Nanobody80 Loop Peptidomimetics: Towards Biased β2 Adrenergic Receptor Ligands

Charlotte Martin; Samuel L C Moors; Mia Danielsen; Cecilia Betti; Cecilia Fabris; Daniel Sejer Pedersen; Els Pardon; Marion Peyressatre; Krisztina Fehér; José C Martins; Jesper Mosolff Mathiesen; May C Morris; Nick Devoogdt; Vicky Caveliers; Frank De Proft; Jan Steyaert; Steven Ballet

doi:10.1002/chem.201701321

Rational Design of Nanobody80 Loop Peptidomimetics: Towards Biased β₂ Adrenergic Receptor Ligands

Charlotte Martin, Samuel L C Moors, Mia Danielsen, Cecilia Betti, Cecilia Fabris, Daniel Sejer Pedersen, Els Pardon, Marion Peyressatre, Krisztina Fehér, José C Martins, Jesper Mosolff Mathiesen, May C Morris, Nick Devoogdt, Vicky Caveliers, Frank De Proft, Jan Steyaert, Steven Ballet

Department of Drug Design and Pharmacology

7 Citations (Scopus)

Abstract

G protein-coupled receptors (GPCRs) play an important role in many cellular responses; as such, their mechanism of action is of utmost interest. To gain insight into the active conformation of GPCRs, the X-ray crystal structures of nanobody (Nb)-stabilized β2 -adrenergic receptor (β2 AR) have been reported. Nb80, in particular, is able to bind the intracellular G protein binding site of β2 AR and stabilize the receptor in an active conformation. Within Nb80, the complementarity-determining region 3 (CDR3) is responsible for most of the binding interactions. Hence, we hypothesized that peptidomimetics of the CDR3 loop might be sufficient for binding to the receptor, inhibiting the interaction of β2 AR with intracellular GPCR interacting proteins (e.g., G proteins). Based on previous crystallographic data, a set of peptidomimetics were synthesized that, similar to the Nb80 CDR3 loop, adopt a β-hairpin conformation. Syntheses, conformational analysis, binding and functional in vitro assays, as well as internalization experiments, were performed. We demonstrate that peptidomimetics can structurally mimic the CDR3 loop of a nanobody and its function by inhibiting G protein coupling as measured by partial inhibition of cAMP production.

Original language	English
Journal	Chemistry: A European Journal
Volume	23
Issue number	40
Pages (from-to)	9632-9640
Number of pages	9
ISSN	0947-6539
DOIs	https://doi.org/10.1002/chem.201701321
Publication status	Published - 18 Jul 2017

Keywords

Journal Article

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10.1002/chem.201701321

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Martin, C., Moors, S. L. C., Danielsen, M., Betti, C., Fabris, C., Sejer Pedersen, D., Pardon, E., Peyressatre, M., Fehér, K., Martins, J. C., Mosolff Mathiesen, J., Morris, M. C., Devoogdt, N., Caveliers, V., De Proft, F., Steyaert, J., & Ballet, S. (2017). Rational Design of Nanobody80 Loop Peptidomimetics: Towards Biased β₂ Adrenergic Receptor Ligands. Chemistry: A European Journal, 23(40), 9632-9640. https://doi.org/10.1002/chem.201701321

Martin, C, Moors, SLC, Danielsen, M, Betti, C, Fabris, C, Sejer Pedersen, D, Pardon, E, Peyressatre, M, Fehér, K, Martins, JC, Mosolff Mathiesen, J, Morris, MC, Devoogdt, N, Caveliers, V, De Proft, F, Steyaert, J & Ballet, S 2017, 'Rational Design of Nanobody80 Loop Peptidomimetics: Towards Biased β₂ Adrenergic Receptor Ligands', Chemistry: A European Journal, vol. 23, no. 40, pp. 9632-9640. https://doi.org/10.1002/chem.201701321

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abstract = "G protein-coupled receptors (GPCRs) play an important role in many cellular responses; as such, their mechanism of action is of utmost interest. To gain insight into the active conformation of GPCRs, the X-ray crystal structures of nanobody (Nb)-stabilized β2 -adrenergic receptor (β2 AR) have been reported. Nb80, in particular, is able to bind the intracellular G protein binding site of β2 AR and stabilize the receptor in an active conformation. Within Nb80, the complementarity-determining region 3 (CDR3) is responsible for most of the binding interactions. Hence, we hypothesized that peptidomimetics of the CDR3 loop might be sufficient for binding to the receptor, inhibiting the interaction of β2 AR with intracellular GPCR interacting proteins (e.g., G proteins). Based on previous crystallographic data, a set of peptidomimetics were synthesized that, similar to the Nb80 CDR3 loop, adopt a β-hairpin conformation. Syntheses, conformational analysis, binding and functional in vitro assays, as well as internalization experiments, were performed. We demonstrate that peptidomimetics can structurally mimic the CDR3 loop of a nanobody and its function by inhibiting G protein coupling as measured by partial inhibition of cAMP production.",

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T2 - Towards Biased β2 Adrenergic Receptor Ligands

AU - Martin, Charlotte

AU - Moors, Samuel L C

AU - Danielsen, Mia

AU - Betti, Cecilia

AU - Fabris, Cecilia

AU - Sejer Pedersen, Daniel

AU - Pardon, Els

AU - Peyressatre, Marion

AU - Fehér, Krisztina

AU - Martins, José C

AU - Mosolff Mathiesen, Jesper

AU - Morris, May C

AU - Devoogdt, Nick

AU - Caveliers, Vicky

AU - De Proft, Frank

AU - Steyaert, Jan

AU - Ballet, Steven

PY - 2017/7/18

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N2 - G protein-coupled receptors (GPCRs) play an important role in many cellular responses; as such, their mechanism of action is of utmost interest. To gain insight into the active conformation of GPCRs, the X-ray crystal structures of nanobody (Nb)-stabilized β2 -adrenergic receptor (β2 AR) have been reported. Nb80, in particular, is able to bind the intracellular G protein binding site of β2 AR and stabilize the receptor in an active conformation. Within Nb80, the complementarity-determining region 3 (CDR3) is responsible for most of the binding interactions. Hence, we hypothesized that peptidomimetics of the CDR3 loop might be sufficient for binding to the receptor, inhibiting the interaction of β2 AR with intracellular GPCR interacting proteins (e.g., G proteins). Based on previous crystallographic data, a set of peptidomimetics were synthesized that, similar to the Nb80 CDR3 loop, adopt a β-hairpin conformation. Syntheses, conformational analysis, binding and functional in vitro assays, as well as internalization experiments, were performed. We demonstrate that peptidomimetics can structurally mimic the CDR3 loop of a nanobody and its function by inhibiting G protein coupling as measured by partial inhibition of cAMP production.

AB - G protein-coupled receptors (GPCRs) play an important role in many cellular responses; as such, their mechanism of action is of utmost interest. To gain insight into the active conformation of GPCRs, the X-ray crystal structures of nanobody (Nb)-stabilized β2 -adrenergic receptor (β2 AR) have been reported. Nb80, in particular, is able to bind the intracellular G protein binding site of β2 AR and stabilize the receptor in an active conformation. Within Nb80, the complementarity-determining region 3 (CDR3) is responsible for most of the binding interactions. Hence, we hypothesized that peptidomimetics of the CDR3 loop might be sufficient for binding to the receptor, inhibiting the interaction of β2 AR with intracellular GPCR interacting proteins (e.g., G proteins). Based on previous crystallographic data, a set of peptidomimetics were synthesized that, similar to the Nb80 CDR3 loop, adopt a β-hairpin conformation. Syntheses, conformational analysis, binding and functional in vitro assays, as well as internalization experiments, were performed. We demonstrate that peptidomimetics can structurally mimic the CDR3 loop of a nanobody and its function by inhibiting G protein coupling as measured by partial inhibition of cAMP production.

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