Structural analysis of rhodopsin states in megabody complexes
Data files
Mar 09, 2026 version files 319.03 KB
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Fig_1.xlsx
175.83 KB
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Fig_2.xlsx
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README.md
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Abstract
Bovine rhodopsin (bRho), the most intensively studied G protein–coupled receptor (GPCR), is activated by light-induced isomerization of its chromophore 11-cis-retinal to all-trans-retinal. This study employed cryogenic electron microscopy (cryo-EM) to investigate rhodopsin structure using a megabody (Mb7) as a negative allosteric modulator.
The data shows the chromatographic purification of Mb7 in complex with bRho, and Mb7 in complex with apo-rhodopsin (Opsin). In addition, we show UV-visible absorption spectra, together with time course of A470 nm and fluorescence emission, that demonstrate that Mb7 acts as a negative allosteric modulator by shifting the equilibrium of photoactivated rhodopsin (bRho*) from the active Meta-II conformation to the inactive Meta-I state.
Dataset DOI: 10.5061/dryad.pg4f4qs4d
Description of the data and file structure
This data comprises purification and spectroscopic characterization efforts for the complexes between a megabody (Mb7) and ground-state rhodopsin (Rho), photoactivated rhodopsin (Rho*) and apo-rhodopsin (Opsin). Empty cells in the dataset are intentional and do not represent missing data.
Files and variables
File: Fig_1.xlsx
Description: Purification of Rho/Mb7 and Opsin/Mb7 (Fig. 1)
Variables
- Column A: Fig. 1A (x). Elution volume (mL) (Rho/Mb7 chromatogram)
- Column B: Fig. 1A (y). A280 nm (Rho/Mb7 chromatogram)
- Column C: Fig. 1C (x). Elution volume (mL) (Opsin/Mb7 chromatogram)
- Column D: Fig. 1C (y). A280 nm (Opsin/Mb7 chromatogram)
- Column E: Fig. 1A inset (x). Wavelength (nm) (Rho/Mb7)
- Column F: Fig. 1A inset (y). Absorbance (Rho/Mb7)
- Column G: Fig. 1C inset (x). Wavelength (nm) (Opsin/Mb7)
- Column H: Fig. 1C inset (y). Absorbance (Opsin/Mb7)
File: Fig_2.xlsx
Description: Spectroscopty of Mb7/rhodopsin complexes (Fig. 2)
Variables
- Column A: Fig. 2A (x). Wavelength (nm)
- Column B: Fig. 2A (y1). Absorbance (bRho)
- Column C: Fig. 2A (y2). Absorbance (bRho*)
- Column D: Fig. 2B (x). Wavelength (nm)
- Column E: Fig. 2B (y1). bRho* (1h, RT)
- Column F: Fig. 2B (y2). Mb7/bRho* (1 h, RT)
- Column G: Fig. 2B (y3). bRho* (20.5 h, ice)
- Column H: Fig. 2B (y4). Mb7/bRho* (20.5 h, ice)
- Column I: Fig. 2C (x). Wavelength (nm)
- Column J: Fig. 2C (y1). Absorbance (bRho/Mb7)
- Column K: Fig. 2C (y2). Absorbance (bRho*/Mb7)
- Column L: Fig. 2D (x1). Time (h) (bRho*, RT)
- Column M: Fig. 2D (y1). A470 nm (bRho*, RT)
- Column N: Fig. 2D (x2). Time (h) (bRho*/Mb7, RT)
- Column O: Fig. 2D (y2). A470 nm (bRho*/Mb7, RT)
- Column P: Fig. 2D (x3). Time (h) (bRho*, ice)
- Column Q: Fig. 2D (y3). A470 nm (bRho*, ice)
- Column R: Fig. 2D (x4). Time (h) (bRho*/Mb7, ice)
- Column S: Fig. 2D (y4). A470 nm (bRho*/Mb7, ice)
- Column T: Fig. 2E (y). Time (s)
- Column U: Fig. 2E (x1). No Mb7 (control)
- Column V: Fig. 2E (x2). [Mb7]= 420 nM
- Column W: Fig. 2E (x3). [Mb7]= 600 nM
- Column X: Fig. 2E (x4). [Mb7]= 840 nM