Light-induced interaction between rhodopsin and GTP-binding protein leads to the hydrolysis of GTP in the rod outer segment

1986 ◽  
Vol 64 (4) ◽  
pp. 304-308 ◽  
Author(s):  
B. D. Gupta ◽  
T. J. Borys ◽  
S. Deshpande ◽  
R. E. Jones ◽  
E. W. Abrahamson
Keyword(s):  
Light Scattering ◽  
Outer Segment ◽  
Plasma Membranes ◽  
Binding Protein ◽  
Rod Outer Segments ◽  
Particle Scattering ◽  
Gtp Binding Protein ◽  
Gtp Binding ◽  
Time Period ◽  
Hydrolysis Of

In the presence of exogeneous GTP, vertebrate whole rod outer segments (ROS), with perforated plasma membranes in the "single particle" scattering range, elicit a light-induced light-scattering transient which we call the "G" signal. Here, we report on the characteristics of the "G" signal relative to the "binding" and "dissociation" signals reported by Kuhn and colleagues. Replacing GTP with guanylyl imidodiphosphate (GMP-PNP) does not give rise to the G signal. This indicates that hydrolysis of the terminal phosphate is required for the G signal and, in addition. GTP and GMP-PNP compete for the same binding site of the enzyme responsible for the G signal (i.e., GTP-binding protein). Also, neither GDP nor its nonhydrolyzable analogue, guanosine 5′-O-(2-thiodiphosphate), when present in ROS suspensions yield any light-scattering transient in the time period tested.

Photoreceptor rod outer segment 48-kDa protein has ATPase activity

1990 ◽  
Vol 5 (6) ◽  
pp. 585-589 ◽  
Author(s):  
Ari Sitaramayya ◽  
Shereen Hakki
Keyword(s):  
Atpase Activity ◽  
Cyclic Gmp ◽  
Slow Rate ◽  
Binding Protein ◽  
Rod Outer Segments ◽  
Light Activation ◽  
Visual Transduction ◽  
Gtp Binding Protein ◽  
Gtp Binding

AbstractThe role of 48-kDa protein in Visual transduction remains unresolved. Two hypotheses for its role in quenching the light activation of cyclic GMP cascade suggest that the protein binds to either phosphodiesterase or phosphorylated rhodopsin. Since the protein is also reported to bind ATP, we anticipated that the protein may have ATP hydrolyzing activity, and in analogy with the GTP-binding protein of the rod outer segments, such activity may be greatly enhanced by the elements of transduction cyclic GMP cascade, permitting the protein to function cyclically as GTP-binding protein does. We found that purified 48-kDa protein hydrolyzes ATP but at a slow rate of 0.04–0.05 per min. The Km for ATP is about 45–65 μM. The activity is inhibited noncompetitively by ADP with a Ki of about 50 μM. The ATPase activity of 48-kDa protein is not affected by rhodopsin, bleached rhodopsin, phosphorylated rhodopsin, unactivated cyclic GMP phosphodiesterase, or phosphodiesterase (PDE) activated by GMP PNP-bound G-protein. These data show that although 48-kDa protein has ATPase activity, lack of regulation of this activity by the elements of visual transduction makes it unlikely for this activity to have a role in quenching the light activation of cyclic GMP cascade.

Localization of ral, a small Mr GTP-binding protein, to collecting duct cells in bovine and rat kidney

1991 ◽  
Vol 261 (6) ◽  
pp. F1063-F1070
Author(s):  
A. Gupta ◽  
B. Bastani ◽  
P. Chardin ◽  
K. A. Hruska
Keyword(s):  
Gene Product ◽  
Plasma Membranes ◽  
Binding Protein ◽  
Collecting Duct ◽  
Rat Kidney ◽  
Kidney Cortex ◽  
Gtp Binding Protein ◽  
Bovine Kidney ◽  
Gtp Binding ◽  
Collecting Ducts

Plasma membranes from bovine kidney cortex were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and transferred to nitrocellulose membranes. Blotting with [alpha-32P]GTP and [35S]GTP gamma S demonstrated specific binding to three and six distinct protein bands, respectively, in the 20,000- to 29,000-Mr range. This indicated the presence of small Mr GTP binding proteins (smg) in bovine kidney cortex. Only one smg with 28,000 Mr was labeled with hydrolysis-resistant GTP photoaffinity probe p3-(4-azidoanilido)-p1-5GTP (AAGTP). The major smg in platelet membranes that binds GTP on nitrocellulose blots has been identified as ral-Mr 29,000. With the use of an antiserum against the ral A gene product, one of the smg with Mr of 29,000 present in bovine renal cortical plasma membranes was identified as ral. Ral was absent from glomerular homogenate, suggesting that it is localized to the tubular segments of the nephron. Ral was detected only in the particulate fraction and not the cytosol. Further subcellular localization of ral was investigated by immunohistochemical staining. Anti-ral antibody immunostained the apical and basolateral membranes of cells in the cortical and medullary collecting ducts in a speckled pattern in the bovine kidney. In the rat kidney, however, uniform linear staining of cortical and medullary collecting ducts predominantly localized to the apical membrane was observed. To date, no function has been assigned to ral. Localization of the ral gene product to the collecting duct suggests a specific functional role for this GTP-binding protein.

scholarly journals Purification and subunit composition of a GTP-binding protein from maize root plasma membranes

FEBS Letters ◽  
1991 ◽  
Vol 291 (2) ◽  
pp. 219-221 ◽  
Author(s):  
Sevo V. Bilushi ◽  
Alexander G. Shebunin ◽  
Alexey V. Babakov
Keyword(s):  
Plasma Membranes ◽  
Binding Protein ◽  
Maize Root ◽  
Subunit Composition ◽  
Gtp Binding Protein ◽  
Gtp Binding

scholarly journals A blue-light-activated GTP-binding protein in the plasma membranes of etiolated peas.

1991 ◽  
Vol 88 (20) ◽  
pp. 8925-8929 ◽  
Author(s):  
K. M. Warpeha ◽  
H. E. Hamm ◽  
M. M. Rasenick ◽  
L. S. Kaufman
Keyword(s):  
Blue Light ◽  
Plasma Membranes ◽  
Binding Protein ◽  
Gtp Binding Protein ◽  
Gtp Binding
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