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Proton Transfer and Protein Conformation Dynamics in Photosensitive Proteins by Time-resolved Step-scan Fourier-transform Infrared Spectroscopy
Published on: June 27, 2014
Two protonation switches control rhodopsin activation in membranes
Mohana Mahalingam1, Karina Martínez-Mayorga, Michael F Brown
1Biophysics Section, Institute of Molecular Medicine and Cell Research, Albert Ludwigs University, Hermann-Herder-Strasse 9, D-79104 Freiburg, Germany.
Summary
Light activates rhodopsin through two protonation switches. At physiological temperatures, these switches partially uncouple, creating an intermediate state. Full activation requires proton uptake by Glu-134, crucial for GPCR signaling.
Area of Science:
- Biochemistry
- Molecular Biology
- Biophysics
Background:
- G protein-coupled receptor (GPCR) activation, exemplified by rhodopsin, involves light-induced retinal isomerization.
- This triggers conformational changes, including two critical protonation switches, leading to the active Meta II state.
- The first switch breaks a salt bridge via proton transfer from the retinal protonated Schiff base (PSB) to Glu-113; the second involves Glu-134 protonation from the solvent.
Purpose of the Study:
- To investigate the activation mechanism of rhodopsin in various membrane environments.
- To elucidate the interplay and coupling of the two protonation switches during receptor activation.
- To determine the role of Glu-134 protonation in stabilizing the active receptor conformation.
Main Methods:
- UV-visible spectroscopy
- Fourier-transform infrared (FTIR) spectroscopy
- Studies in different membrane environments
Main Results:
- At physiological temperatures, the two protonation switches become partially uncoupled.
- A stable, entropy-stabilized intermediate (Meta II) is formed with the salt bridge broken but Glu-134 unprotonated.
- Full activation to the stable Meta II state requires pH-dependent protonation of Glu-134, which is thermodynamically stabilizing.
Conclusions:
- Protonation of Glu-134 is a thermodynamic requirement for full rhodopsin activation, not solely a structural prerequisite.
- The conserved E(D)RY motif's carboxylate protonation likely plays a similar role in signaling for other GPCRs.
- Understanding these protonation dynamics is key to GPCR signal transduction mechanisms.
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