Transducin‐mediated, isoform‐specific interaction of recombinant rat nucleoside diphosphate kinases with bleached bovine retinal rod outer segment membranes

Orlov, N. Yu.; Orlova, T. G.; Nomura, Koji; Hanai, Nobuo; Kimura, Narimichi

doi:10.1016/0014-5793(96)00575-3

Cited by 24 publications

(15 citation statements)

References 23 publications

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“…Furthermore, the effect was specific for the NDPK B isoform. Orlov et al (25) recently found a transducin-mediated isoformspecific binding of NDPK to rod outer segment membranes. In their study, the transducin-mediated binding of NDPK B to the membrane exceeded that for the A isoform 100-fold, indicating a specific interaction of the G protein with NDPK B.…”

Section: Discussionmentioning

confidence: 99%

Activation of Heterotrimeric G Proteins by a High Energy Phosphate Transfer via Nucleoside Diphosphate Kinase (NDPK) B and Gβ Subunits

Hippe¹,

Lutz²,

Cuello³

et al. 2003

Journal of Biological Chemistry

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Formation of GTP by nucleoside diphosphate kinase (NDPK) can contribute to G protein activation in vitro.To study the effect of NDPK on G protein activity in living cells, the NDPK isoforms A and B were stably expressed in H10 cells, a cell line derived from neonatal rat cardiomyocytes. Overexpression of either NDPK isoform had no effect on cellular GTP and ATP levels, basal cAMP levels, basal adenylyl cyclase activity, and the expression of G s ␣ and G i ␣ proteins. However, co-expression of G s ␣ led to an increase in cAMP synthesis that was largely enhanced by the expression of NDPK B, but not NDPK A, and that was confirmed by direct measurement of adenylyl cyclase activity. Cells expressing an inactive NDPK B mutant (H118N) exhibited a decreased cAMP formation in response to G s ␣. Co-immunoprecipitation studies demonstrated a complex formation of the NDPK with G␤␥ dimers. The overexpression of NDPK B, but not its inactive mutant or NDPK A, increased the phosphorylation of G␤ subunits. In summary, our data demonstrate a specific NDPK B-mediated activation of a G protein in intact cells, which is apparently caused by formation of NDPK B⅐G␤␥ complexes and which appears to contribute to the receptor-independent activation of heterotrimeric G proteins. Nucleoside diphosphate kinase (NDPK)1 catalyzes the transfer of terminal phosphate groups from 5Ј-triphosphate to 5Ј-diphosphate nucleotides. In the cell, the major reaction is the phosphate transfer from ATP to other NDPs to maintain the levels of NTPs, especially the relatively high level of GTP. Only a small fraction of cellular NDPK binds to the plasma membrane, where it may serve the synthesis of GTP, required for the activation of G proteins (1-3). An activation of G proteins by NDPK has been disputed for more than 10 years. Although numerous in vitro studies (4 -7) have shown G protein activation through the enzymatic activity of NDPK (synthesis of GTP from a nucleoside triphosphate and GDP), the specificity of this phenomenon has been questioned (8, 9). Particularly in the intact cell, where GTP concentrations are in the upper micromolar range, evidence for a mechanism beyond the sole synthesis of GTP appears mandatory to support this hypothesis. On the other hand, we have shown recently (10) that NDPK activates G proteins and regulates adenylyl cyclase activity in canine cardiac sarcolemmal membranes. This activation required the catalytic activity of NDPK (synthesis of GTP) but was clearly distinct from the effect of exogenous GTP, suggesting a more direct interaction of NDPK and G proteins.Evaluation of direct G protein activation through phosphotransfer by NDPK is associated with substantial methodological constraints. Mainly, GDP is released spontaneously from G proteins and may then serve as a free substrate for phosphorylation by the NDPK (8). Approaches to immobilize the bound GDP at the G protein (11) are associated with protein denaturation, which in turn may lead to unspecific protein phosphorylation by the NDPK (12). In addition, structural con...

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Section: Discussionmentioning

confidence: 99%

Activation of Heterotrimeric G Proteins by a High Energy Phosphate Transfer via Nucleoside Diphosphate Kinase (NDPK) B and Gβ Subunits

Hippe¹,

Lutz²,

Cuello³

et al. 2003

Journal of Biological Chemistry

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“…In the present work the attempt was made to clear up the mechanism of the pH and salt effects on the interaction between R*-G t and NDP kinase. For this purpose, we used the recombinant rat NDP kinase c~ (NDP kinase c~) that strongly interacts with ROS membranes (14). During the first step of this work it was shown that the pH and salt effects on the binding of NDP kinase c~ to the bovine R*-G t containing ROS membranes practically coincide with those measured earlier for the ROS NDP kinase (11,12).…”

Section: Introductionmentioning

confidence: 94%

“…Preparation of NDP kinase a. NDP kinase c~ was prepared and stored as described earlier ( 14,16). Molar extinction coefficients of hexameric NDP kinase c~ (M,.…”

Section: Materials'mentioning

confidence: 99%

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Interaction of recombinant rat nucleoside diphosphate kinase α with bleached bovine retinal rod outer segment membranes: A possible mode of pH and salt effects

et al. 1997

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SummaryAn attempt was made to reveal the mode of action of protons and salts on the recently discovered GTPyS-dependent interaction of bovine retinal rod outer segments (ROS) l nucleoside diphosphate kinase (NDP kinase) with the complex between bleached visual receptor rhodopsin and retinal G-protein transducin in bovine ROS membranes. The properties of recombinant rat NDP kinase cq that is immunologically similar to the soluble NDP kinase from bovine ROS preparation, have been studied in solution by means of protein fluorescence at different pH and salt concentrations and results were compared with pH and salt effects on the binding of NDP kinase a to bleached bovine ROS membranes. The results suggest that NDP kinase c~ itself may serve as a target for protons and salts and mediates their effects on the interaction between the enzyme and ROS membranes.

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“…a complex formation of at least a monomeric NDPK molecule with the heterotrimeric G protein or one of its subunits occurs. Indeed, experimental data have been reported already in the late 1980s, which support such a complex of membranous NDPK with the stimulatory G protein of adenylyl cyclase (G s ; Kimura andShimada 1988a,b, 1990) or the retinal G protein transducin (G t ; Orlov et al 1996;Klinker and Seifert 1999). Nevertheless, the contribution of local GTP formation to effector regulation was disputed or not detected in other systems (Xu et al 1996;Sorota et al 1998).…”

Section: Introductionmentioning

confidence: 99%

Interaction of nucleoside diphosphate kinase B with heterotrimeric G protein βγ dimers: consequences on G protein activation and stability

Wieland

2007

Naunyn-Schmied Arch Pharmacol

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Transducin‐mediated, isoform‐specific interaction of recombinant rat nucleoside diphosphate kinases with bleached bovine retinal rod outer segment membranes

Cited by 24 publications

References 23 publications

Activation of Heterotrimeric G Proteins by a High Energy Phosphate Transfer via Nucleoside Diphosphate Kinase (NDPK) B and Gβ Subunits

Activation of Heterotrimeric G Proteins by a High Energy Phosphate Transfer via Nucleoside Diphosphate Kinase (NDPK) B and Gβ Subunits

Interaction of recombinant rat nucleoside diphosphate kinase α with bleached bovine retinal rod outer segment membranes: A possible mode of pH and salt effects

Interaction of nucleoside diphosphate kinase B with heterotrimeric G protein βγ dimers: consequences on G protein activation and stability

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