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Structural information and membrane binding of truncated RGS9-1 Anchor Protein and its C-terminal hydrophobic segment.

Authors
  • Bernier, Sarah C1
  • Millette, Marc-Antoine1
  • Roy, Sarah1
  • Cantin, Line1
  • Coutinho, Ana2
  • Salesse, Christian3
  • 1 CUO-Recherche, Centre de recherche du CHU de Québec and Département d'ophtalmologie, Faculté de Médecine, and Regroupement Stratégique PROTEO, Université Laval, Québec, Québec, Canada. , (Canada)
  • 2 iBB-Institute for Bioengineering and Biosciences, Department of Bioengineering, Instituto Superior Técnico, Universidade de Lisboa, Av. Rovisco Pais, 1049-001 Lisboa, Portugal; Department of Chemistry and Biochemistry, Faculdade de Ciências, Universidade de Lisboa, Campo Grande, 1749-016 Lisboa, Portugal. , (Portugal)
  • 3 CUO-Recherche, Centre de recherche du CHU de Québec and Département d'ophtalmologie, Faculté de Médecine, and Regroupement Stratégique PROTEO, Université Laval, Québec, Québec, Canada. Electronic address: [email protected] , (Canada)
Type
Published Article
Journal
Biochimica et biophysica acta. Biomembranes
Publication Date
Jan 13, 2021
Volume
1863
Issue
4
Pages
183566–183566
Identifiers
DOI: 10.1016/j.bbamem.2021.183566
PMID: 33453187
Source
Medline
Keywords
Language
English
License
Unknown

Abstract

Visual phototransduction takes place in photoreceptor cells. Light absorption by rhodopsin leads to the activation of transducin as a result of the exchange of its GDP for GTP. The GTP-bound ⍺-subunit of transducin then activates phosphodiesterase (PDE), which in turn hydrolyzes cGMP leading to photoreceptor hyperpolarization. Photoreceptors return to the dark state upon inactivation of these proteins. In particular, PDE is inactivated by the protein complex R9AP/RGS9-1/Gβ5. R9AP (RGS9-1 anchor protein) is responsible for the membrane anchoring of this protein complex to photoreceptor outer segment disk membranes most likely by the combined involvement of its C-terminal hydrophobic domain as well as other types of interactions. This study thus aimed to gather information on the structure and membrane binding of the C-terminal hydrophobic segment of R9AP as well as of truncated R9AP (without its C-terminal domain, R9AP∆TM). Circular dichroism and infrared spectroscopic measurements revealed that the secondary structure of R9AP∆TM mainly includes ⍺-helical structural elements. Moreover, intrinsic fluorescence measurements of native R9AP∆TM and individual mutants lacking one tryptophan demonstrated that W79 is more buried than W173 but that they are both located in a hydrophobic environment. This method also revealed that membrane binding of R9AP∆TM does not involve regions near its tryptophan residues, while infrared spectroscopy validated its binding to lipid vesicles. Additional fluorescence measurements showed that the C-terminal segment of R9AP is membrane embedded. Maximum insertion pressure and synergy data using Langmuir monolayers suggest that interactions with specific phospholipids could be involved in the membrane binding of R9AP∆TM. Copyright © 2021 Elsevier B.V. All rights reserved.

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