Rhodopsin mobility, structure, and lipid-protein interaction in squid photoreceptor membranes

Treatment of outer segment membranes from Loligo forbesi with endoprotease-V8 from Staphylococcus aureus results in cleavage of the C-terminal extension of the squid rhodopsin, with accompanying reduction of the apparent molecular weight from 47,000 to 36,000 on sodium dodecyl sulfate/polyacrylamide...

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Bibliographic Details
Published in:	Biochemistry (Easton) Vol. 32; no. 13; pp. 3298 - 3305
Main Authors:	Ryba, Nicholas J. P, Hoon, Mark A, Findlay, John B. C, Saibil, Helen R, Wilkinson, J. Richard, Heimburg, Thomas, Marsh, Derek
Format:	Journal Article
Language:	English
Published:	Washington, DC American Chemical Society 01-04-1993
Subjects:	Amides Animals Biological and medical sciences Cattle Cell Membrane - chemistry cell membranes Cell physiology Cytoplasm - ultrastructure Decapodiformes Electron Spin Resonance Spectroscopy Fundamental and applied biological sciences. Psychology In Vitro Techniques interaction lipids Loligo forbesi Marine marine molluscs Microscopy, Electron mobility Molecular and cellular biology Motion outer membranes Phosphatidylcholines Photoreceptor Cells - chemistry photoreceptors proteins rhodopsin Rhodopsin - chemistry Rod Cell Outer Segment - chemistry Rod Cell Outer Segment - ultrastructure Spectrophotometry, Infrared Spin Labels structure Structure-Activity Relationship Vision, photoreception Proteins Cephalopoda Rhodopsin Lipids Molecular interaction Invertebrata Mollusca Photoprotein
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Summary:	Treatment of outer segment membranes from Loligo forbesi with endoprotease-V8 from Staphylococcus aureus results in cleavage of the C-terminal extension of the squid rhodopsin, with accompanying reduction of the apparent molecular weight from 47,000 to 36,000 on sodium dodecyl sulfate/polyacrylamide gel electrophoresis. Negative-stain electron microscopy of the intact membranes shows that small clusters of the rhodopsin C-termini form structures extending from the membrane surface and that these are absent after protease treatment. Fourier transform infrared spectra of the amide I band of the protein indicate that removal of the C-terminal extension increases the relative alpha-helical content of squid rhodopsin to a level comparable to that for bovine rhodopsin in disk membranes, and to an extent which suggests that the alpha-helical structure lies mainly in the M(r) 36,000 (transmembrane) section of the protein. Saturation-transfer electron spin resonance (ESR) spectroscopy of the spin-labeled protein reveals that the rotational diffusion of squid rhodopsin in outer segment membranes that have been extensively washed with urea to remove peripheral proteins is much slower than that of bovine rhodopsin in rod outer segment disk membranes. This reduction in rotational mobility is also found with purified squid rhodopsin reconstituted in egg phosphatidylcholine and in urea-washed outer segment membranes which have been treated with endoprotease-V8 to remove the C-terminal extension of squid rhodopsin. In the latter case, the saturation-transfer ESR spectra are virtually identical to those of the non-proteolyzed membranes.
Bibliography:	ark:/67375/TPS-XBRFZ3LN-1 istex:867BA7EDB05348E485E627EBCEB086F6ED77F2E8 ObjectType-Article-1 SourceType-Scholarly Journals-1 ObjectType-Feature-2 content type line 23
ISSN:	0006-2960 1520-4995
DOI:	10.1021/bi00064a012