Current address: Craig McKibbin, Faculty of Life Sciences, Michael Smith Building, Oxford Road, University of Manchester, Manchester M13 9PT, UK.
Urea Unfolding of Opsin in Phospholipid Bicelles†
Article first published online: 13 JAN 2009
© 2009 The Authors. Journal Compilation. The American Society of Photobiology
Photochemistry and Photobiology
Volume 85, Issue 2, pages 494–500, March/April 2009
How to Cite
McKibbin, C., Farmer, N. A., Edwards, P. C., Villa, C. and Booth, P. J. (2009), Urea Unfolding of Opsin in Phospholipid Bicelles. Photochemistry and Photobiology, 85: 494–500. doi: 10.1111/j.1751-1097.2008.00503.x
This paper is part of the Proceedings of the 13th International Conference on Retinal Proteins, Barcelona, Spain, 15–19 June 2008.
- Issue published online: 25 FEB 2009
- Article first published online: 13 JAN 2009
- Received 3 July 2008, accepted 14 October 2008
Opsin is the unstable apo-protein of the light-activated G protein-coupled receptor rhodopsin. We investigated the stability of bovine opsin, solubilized in 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC)/detergent bicelles, against urea-induced unfolding. A single irreversible protein unfolding transition was observed from changes in intrinsic tryptophan fluorescence and far-UV circular dichroism. This unfolding transition correlated with loss of protein activity. Changes in tertiary structure, as indicated by fluorescence measurements, were concomitant with an approximate 50% reduction in α-helical content of opsin, indicating that global unfolding had been induced by urea. The urea concentration at the midpoint of unfolding was dependent on the lipid/detergent environment, occurring at approximately 1.2 m urea in DMPC/1,2-dihexanoyl-sn-glycero-3-phosphocholine bicelles, while being significantly stabilized to approximately 3.5 m urea in DMPC/3-[(cholamidopropyl)dimethylammonio]-1-propanesulfonate bicelles. These findings demonstrate that interactions with the surrounding lipids and detergent are highly influential in the unfolding of membrane protein structure. The urea/bicelle system offers the possibility for a more detailed understanding of the structural changes that take place upon irreversible unfolding of opsin.