NDLI: Aggregation of frog rhodopsin to oligomers and their dissociation to monomer: Application of BN- and SDS-PAGE

Content Provider	Springer Nature Link
Author	Shukolyukov, S. A.
Copyright Year	2009
Abstract	After solubilization of frog rod outer segments (ROS) with mild detergents (digitonin, n-dodecyl-β-D-maltoside, Chaps, Triton X-100) and subsequent one-dimensional blue native polyacrylamide gel electrophoresis (1D BN-PAGE), the position of rhodopsin (Rh) on the gradient gel does not match the monomer with molecular weight of 40 kDa but appears self-associated into aggregate of Rh (RhA) with molecular mass varying in different detergents from 85 to 125 kDa. Short-term treatment (∼2 h) of the excised BN-PAGE strip containing RhA by denaturing detergent mixture (10% SDS + 1 mM dithiothreitol (DTT)) followed by 2D SDS-PAGE revealed dissociation of the RhA into opsin monomer and unidentified proteins. Long-term treatment (∼2 days) of RhA that included extraction, denaturation, concentration, and electrophoresis induced, along with dissociation of RhA into opsin monomer + unidentified proteins, also formation of opsin dimers, trimers, and higher oligomers owing to a secondary aggregation of opsin. Direct solubilization of the ROS by harsh SDS + DTT detergent mixture followed by 1D SDS-PAGE revealed only opsin monomer that upon heating disappeared, trans-forming into higher oligomers owing to secondary aggregation. The data show that degree of Rh oligomerization depends on specific conditions in which it stays. In the native state in the photoreceptor membrane as well as in mild detergents frog Rh exists mainly as dimers or higher oligomers. After solubilization with denaturing detergents, RhA can dissociate into monomers that then spontaneously self-associate into higher oligomers under the influence of various factors (for example, heating).
Starting Page	599
Ending Page	604
Page Count	6
File Format	PDF
ISSN	00062979
Journal	Biochemistry (Moscow)
Volume Number	74
Issue Number	6
e-ISSN	16083040
Language	English
Publisher	SP MAIK Nauka/Interperiodica
Publisher Date	2009-06-19
Publisher Place	Dordrecht
Access Restriction	One Nation One Subscription (ONOS)
Subject Keyword	frog rhodopsin oligomerization dissociation BN-PAGE SDS-PAGE Biomedicine general Microbiology Bioorganic Chemistry Biochemistry
Content Type	Text
Resource Type	Article
Subject	Biochemistry Biophysics Geriatrics and Gerontology

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