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Differential Dynamics of Extracellular and Cytoplasmic Domains in Denatured States of Rhodopsin

[Image: see text] Rhodopsin is a model system for understanding membrane protein folding. Recently, conditions that allow maximally denaturing rhodopsin without causing aggregation have been determined, opening the door to the first structural characterization of denatured states of rhodopsin by nuc...

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Autores principales: Dutta, Arpana, Altenbach, Christian, Mangahas, Sheryll, Yanamala, Naveena, Gardner, Eric, Hubbell, Wayne L., Klein-Seetharaman, Judith
Formato: Online Artículo Texto
Lenguaje:English
Publicado: American Chemical Society 2014
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4245987/
https://www.ncbi.nlm.nih.gov/pubmed/25268658
http://dx.doi.org/10.1021/bi401557e
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author Dutta, Arpana
Altenbach, Christian
Mangahas, Sheryll
Yanamala, Naveena
Gardner, Eric
Hubbell, Wayne L.
Klein-Seetharaman, Judith
author_facet Dutta, Arpana
Altenbach, Christian
Mangahas, Sheryll
Yanamala, Naveena
Gardner, Eric
Hubbell, Wayne L.
Klein-Seetharaman, Judith
author_sort Dutta, Arpana
collection PubMed
description [Image: see text] Rhodopsin is a model system for understanding membrane protein folding. Recently, conditions that allow maximally denaturing rhodopsin without causing aggregation have been determined, opening the door to the first structural characterization of denatured states of rhodopsin by nuclear magnetic resonance (NMR) and electron paramagnetic resonance (EPR) spectroscopy. One-dimensional (1)H NMR spectra confirm a progressive increase in flexibility of resonances in rhodopsin with increasing denaturant concentrations. Two-dimensional (1)H–(15)N HSQC spectra of [(15)N]-α-lysine-labeled rhodopsin in which signals arise primarily from residues in the cytoplasmic (CP) domain and of [(15)N]-α,ε-tryptophan-labeled rhodopsin in which signals arise only from transmembrane (TM) and extracellular (EC) residues indicate qualitatively that EC and CP domains may be differentially affected by denaturation. To obtain residue-specific information, particular residues in EC and CP domains were investigated by site-directed spin labeling. EPR spectra of the spin-labeled samples indicate that the EC residues retain more rigidity in the denatured states than the CP residues. These results support the notion of residual structure in denatured states of rhodopsin.
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spelling pubmed-42459872015-09-30 Differential Dynamics of Extracellular and Cytoplasmic Domains in Denatured States of Rhodopsin Dutta, Arpana Altenbach, Christian Mangahas, Sheryll Yanamala, Naveena Gardner, Eric Hubbell, Wayne L. Klein-Seetharaman, Judith Biochemistry [Image: see text] Rhodopsin is a model system for understanding membrane protein folding. Recently, conditions that allow maximally denaturing rhodopsin without causing aggregation have been determined, opening the door to the first structural characterization of denatured states of rhodopsin by nuclear magnetic resonance (NMR) and electron paramagnetic resonance (EPR) spectroscopy. One-dimensional (1)H NMR spectra confirm a progressive increase in flexibility of resonances in rhodopsin with increasing denaturant concentrations. Two-dimensional (1)H–(15)N HSQC spectra of [(15)N]-α-lysine-labeled rhodopsin in which signals arise primarily from residues in the cytoplasmic (CP) domain and of [(15)N]-α,ε-tryptophan-labeled rhodopsin in which signals arise only from transmembrane (TM) and extracellular (EC) residues indicate qualitatively that EC and CP domains may be differentially affected by denaturation. To obtain residue-specific information, particular residues in EC and CP domains were investigated by site-directed spin labeling. EPR spectra of the spin-labeled samples indicate that the EC residues retain more rigidity in the denatured states than the CP residues. These results support the notion of residual structure in denatured states of rhodopsin. American Chemical Society 2014-09-30 2014-11-25 /pmc/articles/PMC4245987/ /pubmed/25268658 http://dx.doi.org/10.1021/bi401557e Text en Copyright © 2014 American Chemical Society This is an open access article published under an ACS AuthorChoice License (http://pubs.acs.org/page/policy/authorchoice_termsofuse.html) , which permits copying and redistribution of the article or any adaptations for non-commercial purposes.
spellingShingle Dutta, Arpana
Altenbach, Christian
Mangahas, Sheryll
Yanamala, Naveena
Gardner, Eric
Hubbell, Wayne L.
Klein-Seetharaman, Judith
Differential Dynamics of Extracellular and Cytoplasmic Domains in Denatured States of Rhodopsin
title Differential Dynamics of Extracellular and Cytoplasmic Domains in Denatured States of Rhodopsin
title_full Differential Dynamics of Extracellular and Cytoplasmic Domains in Denatured States of Rhodopsin
title_fullStr Differential Dynamics of Extracellular and Cytoplasmic Domains in Denatured States of Rhodopsin
title_full_unstemmed Differential Dynamics of Extracellular and Cytoplasmic Domains in Denatured States of Rhodopsin
title_short Differential Dynamics of Extracellular and Cytoplasmic Domains in Denatured States of Rhodopsin
title_sort differential dynamics of extracellular and cytoplasmic domains in denatured states of rhodopsin
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4245987/
https://www.ncbi.nlm.nih.gov/pubmed/25268658
http://dx.doi.org/10.1021/bi401557e
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