NDLI: Proof of oligomeric state of frog rhodopsin: Visualization of dimer and oligomers on gels after BN- and HRCN-PAGE using antibodies to rhodopsin and by retinylopsin fluorescence

Content Provider	Springer Nature Link
Author	Shukolyukov, S. A.
Copyright Year	2010
Abstract	Staining by antibodies to rhodopsin (Rh) and fluorescence of N-retinylopsin (RO) have shown that digitonin (DIG)-, dodecyl-β-D-maltoside (DM)-, and sodium dodecyl sulfate (SDS)-solubilized frog Rh after BN- and HRCN-PAGE is situated in the gradient gel in the state of dimer with a slight content of higher oligomers (trimer, tetramer, etc.). With increasing detergent harshness (DIG < DM < SDS), the proportion of higher oligomers in extracts becomes more prominent. Formation of RO in rod outer segments (ROS) in the presence of 0.7 M NaBH3CN at pH 5.0 occurs only when Rh is simultaneously photolyzed during reduction. Dithiothreitol at the concentration of 0.005 M failed to induce RO production. Formation of a stable C-N bond between all-trans-retinal and opsin in RO is accompanied by decrease in the dimer share and increase in the share of the higher oligomers due to secondary dissociation-aggregation of solubilized opsin. The position of the Rh dimer in relation to the anode during both native electrophoreses is determined not only by its molecular mass, but probably also depends on unfolding degree (or form): the harsher the detergent, the closer to the anode the dimer is located. Treatment of ROS by agents modifying the cholesterol component of lipid membrane (MβCD, filipin III, nystatin, saponin) did not change the character of Rh oligomerization, thus showing that integrity of the cholesterol component of photoreceptor membrane is not a crucial factor for oligomerization of opsin. It is supposed that the dimer-oligomer “portrait” of frog Rh, which has been found by two methods of native electrophoresis in three detergents with different degree of harshness, corresponds to a physiological state of this protein in native photoreceptor membrane.
Starting Page	1045
Ending Page	1051
Page Count	7
File Format	PDF
ISSN	00062979
Journal	Biochemistry (Moscow)
Volume Number	75
Issue Number	8
e-ISSN	16083040
Language	English
Publisher	SP MAIK Nauka/Interperiodica
Publisher Date	2010-08-29
Publisher Place	Dordrecht
Access Restriction	One Nation One Subscription (ONOS)
Subject Keyword	frog rhodopsin oligomerization BN-PAGE HRCN-PAGE antibodies fluorescence retinylopsin digitonin dodecyl maltoside SDS Biomedicine general Microbiology Bioorganic Chemistry Biochemistry
Content Type	Text
Resource Type	Article
Subject	Biochemistry Biophysics Geriatrics and Gerontology

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