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Nietlispach Group

NMR spectroscopy of membrane proteins

Studying at Cambridge

 

Sensory rhodopsin II

Sensory rhodopsin II (pSRII)

 

Seven-helical membrane proteins represent a challenge for structural biology. To date, only a few sizeable structures of α-helical membrane proteins have been solved by solution NMR. In 2010, our group published the first three-dimensional structure of a seven-helical membrane protein receptor determined by NMR spectroscopy. Sensory rhodopsin II (pSRII), from the archaeon Natronomonas pharaonis, functions via 13-trans–cis isomerization of its all-trans retinal chromophore as a repellent phototactic receptor to blue light, enabling the archaeon to seek the dark when respiratory substrates are plentiful, in order to minimise oxidative damage.

The feasibility of characterising membrane receptors by NMR has opened many new avenues, including studying the folding pathway of such proteins. Our understanding of the folding of membrane proteins lags behind that of soluble proteins due to challenges posed by the exposure of hydrophobic regions during in vitro chemical denaturation and refolding experiments. While different folding models are accepted for soluble proteins, only the two-stage model and the long-range interactions model have been proposed so far for helical membrane proteins. To address our knowledge gap on how different membrane proteins traverse their folding pathways, we are investigating the structural features of the SDS-denatured states and the kinetics for reversible unfolding of pSRII. 

pSRII signals via its downstream transducer, pHtrII, initiating a Histidine kinase/Aspartate kinase signal transduction cascade. This is a paradigm for bacterial signalling systems and we are interested in characterising this interaction and the signalling mechanism using NMR and other techniques such as cryoEM.

 

 

For more information see:

Structure determination of the seven-helix transmembrane receptor sensory rhodopsin II by solution NMR spectroscopy.
Antoine Gautier, Helen R. Mott, Mark J. Bostock, John P. Kirkpatrick, Daniel Nietlispach, Nature Structural and Molecular Biology (2010), 17, 768-774
(doi:10.1038/nsmb.1807)

and

Solution-state NMR spectroscopy of a seven-helix transmembrane protein receptor: backbone assignment, secondary structure, and dynamics.
Antoine Gautier, John P. Kirkpatrick, Daniel Nietlispach, Angewandte Chemie International Edition (2008), 47, 7297-7300
(doi:10.1002/anie.200802783)