A large geometric distortion in the first photointermediate of rhodopsin, determined by double-quantum solid-state NMR

Concistrè, Maria; Johannessen, Ole G.; McLean, Neville; Bovée-Geurts, Petra H. M.; Brown, Richard C. D.; DeGrip, Willem J.; Levitt, Malcolm H.

doi:10.1007/s10858-012-9635-4

Cited by 10 publications

(13 citation statements)

References 42 publications

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“…For HCCH torsional angle measurements a double quantum heteronuclear local field experiment 64 was applied (HCCH-experiment). In the experiment POST-C7 during two rotor periods was used for double quantum excitation and reconversion, PMLG-9 65…”

Section: Dihedral Angle Determinationmentioning

confidence: 99%

Chromophore Distortions in Photointermediates of Proteorhodopsin Visualized by Dynamic Nuclear Polarization-Enhanced Solid-State NMR

et al. 2017

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Proteorhodopsin (PR) is the most abundant retinal protein on earth and functions as a light-driven proton pump. Despite extensive efforts, structural data for PR photointermediate states have not been obtained. On the basis of dynamic nuclear polarization (DNP)-enhanced solid-state NMR, we were able to analyze the retinal polyene chain between positions C10 and C15 as well as the Schiff base nitrogen in the ground state in comparison to light-induced, cryotrapped K- and M-states. A high M-state population could be achieved by preventing reprotonation of the Schiff base through a mutation of the primary proton donor (E108Q). Our data reveal unexpected large and alternating C chemical shift changes in the K-state propagating away from the Schiff base along the polyene chain. Furthermore, two different M-states have been observed reflecting the Schiff base reorientation after the deprotonation step. Our study provides novel insight into the photocycle of PR and also demonstrates the power of DNP-enhanced solid-state NMR to bridge the gap between functional and structural data and models.

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Section: Dihedral Angle Determinationmentioning

confidence: 99%

Chromophore Distortions in Photointermediates of Proteorhodopsin Visualized by Dynamic Nuclear Polarization-Enhanced Solid-State NMR

et al. 2017

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“…A continuous wave decoupling at 112 kHz and a SPINAL64 (34) decoupling at 105 kHz were applied during POST-C7 pulses and acquisition time, respectively. HCCH torsion angles were measured by separated local field (SLF) experiments, essentially as described by Levitt and co-workers (35): DQ coherences are excited, evolve under homonuclear proton decoupling, and are detected after a reconversion step. Here, two complete POST-C7 cycles were used for both DQ excitation and reconversion.…”

Section: Simulation Of the Transient Datamentioning

confidence: 99%

“…Retinal 13 C chemical shifts are sensitive indicators for conformational changes. In particular, double quantum spectroscopy has been shown to offer unique insight: Chemical shift data can be extracted more easily by natural background suppression and CC bond lengths and HCCH torsion angles can be obtained from DQ built-up curves (40) and SLF experiments (35). Such experiments will also benefit from signal enhancement provided by DNP (29).…”

Section: Solid-state Nmr On Gpr A178rmentioning

confidence: 99%

The EF Loop in Green Proteorhodopsin Affects Conformation and Photocycle dynamics

Mehler

Scholz

Ullrich

et al. 2013

Biophysical Journal

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The proteorhodopsin family consists of retinal proteins of marine bacterial origin with optical properties adjusted to their local environments. For green proteorhodopsin, a highly specific mutation in the EF loop, A178R, has been found to cause a surprisingly large redshift of 20 nm despite its distance from the chromophore. Here, we analyze structural and functional consequences of this EF loop mutation by time-resolved optical spectroscopy and solid-state NMR. We found that the primary photoreaction and the formation of the K-like photo intermediate is almost pH-independent and slower compared to the wild-type, whereas the decay of the K-intermediate is accelerated, suggesting structural changes within the counterion complex upon mutation. The photocycle is significantly elongated mainly due to an enlarged lifetime of late photo intermediates. Multidimensional MAS-NMR reveals mutation-induced chemical shift changes propagating from the EF loop to the chromophore binding pocket, whereas dynamic nuclear polarization-enhanced (13)C-double quantum MAS-NMR has been used to probe directly the retinylidene conformation. Our data show a modified interaction network between chromophore, Schiff base, and counterion complex explaining the altered optical and kinetic properties. In particular, the mutation-induced distorted structure in the EF loop weakens interactions, which help reorienting helix F during the reprotonation step explaining the slower photocycle. These data lead to the conclusion that the EF loop plays an important role in proton uptake from the cytoplasm but our data also reveal a clear interaction pathway between the EF loop and retinal binding pocket, which might be an evolutionary conserved communication pathway in retinal proteins.

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“…Solid-state NMR spectroscopy has been particularly useful in characterizing the structure of dark rhodopsin and its intermediates. Deuterium NMR spectroscopy along with molecular dynamics has provided insights into the structure and dynamics of the retinal chromphore [41], while 13 C and 1 H NMR correlation spectroscopy has been used to probe the retinal structure and its interaction with surrounding amino acids [42, 43] The use of selective pairs of 13 C labels has been useful for characterizing the conformation of the retinal [44] and internuclear 13 C.. 13 C distances in the protein [45]. The receptor structure can be probed in a membrane environment using the native protein sequence or site directed mutants, and low temperature [46] provides a way to trap intermediates.…”

Section: Introductionmentioning

confidence: 99%

Amino acid conservation and interactions in rhodopsin: Probing receptor activation by NMR spectroscopy

Pope

Eilers

Reeves

et al. 2014

Biochimica et Biophysica Acta (BBA) - Bioenergetics

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Rhodopsin is a classical two-state G protein-coupled receptor (GPCR). In the dark, its 11-cis retinal chromophore serves as an inverse agonist to lock the receptor in an inactive state. Retinal-protein and protein-protein interactions have evolved to reduce the basal activity of the receptor in order to achieve low dark noise in the visual system. In contrast, absorption of light triggers rapid isomerization of the retinal, which drives the conversion of the receptor to a fully active conformation. Several specific protein-protein interactions have evolved that maintain the lifetime of the active state in order to increase the sensitivity of this receptor for dim-light vision in vertebrates. In this article, we review the molecular interactions that stabilize rhodopsin in the dark-state and describe the use of solid-state NMR spectroscopy for probing the structural changes that occur upon light-activation. Amino acid conservation provides a guide for those interactions that are common in the class A GPCRs as well as those that are unique to the visual system.

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A large geometric distortion in the first photointermediate of rhodopsin, determined by double-quantum solid-state NMR

Cited by 10 publications

References 42 publications

Chromophore Distortions in Photointermediates of Proteorhodopsin Visualized by Dynamic Nuclear Polarization-Enhanced Solid-State NMR

Chromophore Distortions in Photointermediates of Proteorhodopsin Visualized by Dynamic Nuclear Polarization-Enhanced Solid-State NMR

The EF Loop in Green Proteorhodopsin Affects Conformation and Photocycle dynamics

Amino acid conservation and interactions in rhodopsin: Probing receptor activation by NMR spectroscopy

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