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The role of Gi and the membrane-fluidizing agent benzyl alcohol in modulating the hysteretic activation of human platelet adenylate cyclase by guanylyl 5′-imidodiphosphate
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| Zeitschriftentitel: | Biochemical Journal |
|---|---|
| Personen und Körperschaften: | , |
| In: | Biochemical Journal, 291, 1993, 3, S. 945-949 |
| Format: | E-Article |
| Sprache: | Englisch |
| veröffentlicht: |
Portland Press Ltd.
|
| Schlagwörter: |
| author_facet |
Spence, S Houslay, M D Spence, S Houslay, M D |
|---|---|
| author |
Spence, S Houslay, M D |
| spellingShingle |
Spence, S Houslay, M D Biochemical Journal The role of Gi and the membrane-fluidizing agent benzyl alcohol in modulating the hysteretic activation of human platelet adenylate cyclase by guanylyl 5′-imidodiphosphate Cell Biology Molecular Biology Biochemistry |
| author_sort |
spence, s |
| spelling |
Spence, S Houslay, M D 0264-6021 1470-8728 Portland Press Ltd. Cell Biology Molecular Biology Biochemistry http://dx.doi.org/10.1042/bj2910945 <jats:p>The non-hydrolysable GTP analogue guanylyl 5′-imidodiphosphate (p[NH]ppG) elicited a profound increase in the adenylate cyclase activity of human platelets. This occurred after a well-defined lag period of around 6 min, whereupon an enhanced steady-state rate was evident. The duration of the lag period was unchanged over a range of concentrations of p[NH]ppG which gave very different steady-state rates of adenylate cyclase activity. Prior activation of the stimulatory G-protein Gs by cholera-toxin pre-treatment abolished the lag period and elicited a small increase in the steady-state rate. Manipulating function of the inhibitory G-protein Gi also led to profound changes in the lag periods. Thus marked decreases in the lag were seen (approximately 70-81%) when Gi function was ablated through pre-treatment of platelet membranes with pertussis toxin, or by using elevated (25 mM) Mg2+ levels in the assay, or when Mg2+ was replaced by 5 mM Mn2+ in the assay. In contrast with this, potentiation of Gi function led to an increase in the lag period, as seen under conditions of agonist occupancy of inhibitory alpha 2-adrenoceptors (increase approximately 74%) or with the addition of 100 mM NaCl to the assays (increase approximately 44%). The local anaesthetic and membrane-fluidizing agent benzyl alcohol elicited both a profound decrease (around 70% at 80 mM) in the p[NH]ppG-induced lag period and a marked augmentation (around 5-fold) in the steady-state adenylate cyclase activity. When adenylate cyclase assays were done at 35 degrees C instead of 25 degrees C, then the lag period for activation by p[NH]ppG was decreased by around 33% and the steady-state rate increased by around 3-fold. At 35 degrees C, the addition of benzyl alcohol led to the apparent abolition of the lag period for p[NH]ppG activation of adenylate cyclase and amplified the steady-state rate by only around 2.2-fold. It is shown that Gi plays a fundamental role in determining the rate of activation of Gs. The proposal is formulated that such an action may be mediated through the release of beta gamma-subunits. Thus beta gamma-subunit dissociation is proposed as providing the rate-limiting step in Gi activation.</jats:p> The role of Gi and the membrane-fluidizing agent benzyl alcohol in modulating the hysteretic activation of human platelet adenylate cyclase by guanylyl 5′-imidodiphosphate Biochemical Journal |
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1993 |
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Biochemical Journal |
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49 |
| title |
The role of Gi and the membrane-fluidizing agent benzyl alcohol in modulating the hysteretic activation of human platelet adenylate cyclase by guanylyl 5′-imidodiphosphate |
| title_unstemmed |
The role of Gi and the membrane-fluidizing agent benzyl alcohol in modulating the hysteretic activation of human platelet adenylate cyclase by guanylyl 5′-imidodiphosphate |
| title_full |
The role of Gi and the membrane-fluidizing agent benzyl alcohol in modulating the hysteretic activation of human platelet adenylate cyclase by guanylyl 5′-imidodiphosphate |
| title_fullStr |
The role of Gi and the membrane-fluidizing agent benzyl alcohol in modulating the hysteretic activation of human platelet adenylate cyclase by guanylyl 5′-imidodiphosphate |
| title_full_unstemmed |
The role of Gi and the membrane-fluidizing agent benzyl alcohol in modulating the hysteretic activation of human platelet adenylate cyclase by guanylyl 5′-imidodiphosphate |
| title_short |
The role of Gi and the membrane-fluidizing agent benzyl alcohol in modulating the hysteretic activation of human platelet adenylate cyclase by guanylyl 5′-imidodiphosphate |
| title_sort |
the role of gi and the membrane-fluidizing agent benzyl alcohol in modulating the hysteretic activation of human platelet adenylate cyclase by guanylyl 5′-imidodiphosphate |
| topic |
Cell Biology Molecular Biology Biochemistry |
| url |
http://dx.doi.org/10.1042/bj2910945 |
| publishDate |
1993 |
| physical |
945-949 |
| description |
<jats:p>The non-hydrolysable GTP analogue guanylyl 5′-imidodiphosphate (p[NH]ppG) elicited a profound increase in the adenylate cyclase activity of human platelets. This occurred after a well-defined lag period of around 6 min, whereupon an enhanced steady-state rate was evident. The duration of the lag period was unchanged over a range of concentrations of p[NH]ppG which gave very different steady-state rates of adenylate cyclase activity. Prior activation of the stimulatory G-protein Gs by cholera-toxin pre-treatment abolished the lag period and elicited a small increase in the steady-state rate. Manipulating function of the inhibitory G-protein Gi also led to profound changes in the lag periods. Thus marked decreases in the lag were seen (approximately 70-81%) when Gi function was ablated through pre-treatment of platelet membranes with pertussis toxin, or by using elevated (25 mM) Mg2+ levels in the assay, or when Mg2+ was replaced by 5 mM Mn2+ in the assay. In contrast with this, potentiation of Gi function led to an increase in the lag period, as seen under conditions of agonist occupancy of inhibitory alpha 2-adrenoceptors (increase approximately 74%) or with the addition of 100 mM NaCl to the assays (increase approximately 44%). The local anaesthetic and membrane-fluidizing agent benzyl alcohol elicited both a profound decrease (around 70% at 80 mM) in the p[NH]ppG-induced lag period and a marked augmentation (around 5-fold) in the steady-state adenylate cyclase activity. When adenylate cyclase assays were done at 35 degrees C instead of 25 degrees C, then the lag period for activation by p[NH]ppG was decreased by around 33% and the steady-state rate increased by around 3-fold. At 35 degrees C, the addition of benzyl alcohol led to the apparent abolition of the lag period for p[NH]ppG activation of adenylate cyclase and amplified the steady-state rate by only around 2.2-fold. It is shown that Gi plays a fundamental role in determining the rate of activation of Gs. The proposal is formulated that such an action may be mediated through the release of beta gamma-subunits. Thus beta gamma-subunit dissociation is proposed as providing the rate-limiting step in Gi activation.</jats:p> |
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| description | <jats:p>The non-hydrolysable GTP analogue guanylyl 5′-imidodiphosphate (p[NH]ppG) elicited a profound increase in the adenylate cyclase activity of human platelets. This occurred after a well-defined lag period of around 6 min, whereupon an enhanced steady-state rate was evident. The duration of the lag period was unchanged over a range of concentrations of p[NH]ppG which gave very different steady-state rates of adenylate cyclase activity. Prior activation of the stimulatory G-protein Gs by cholera-toxin pre-treatment abolished the lag period and elicited a small increase in the steady-state rate. Manipulating function of the inhibitory G-protein Gi also led to profound changes in the lag periods. Thus marked decreases in the lag were seen (approximately 70-81%) when Gi function was ablated through pre-treatment of platelet membranes with pertussis toxin, or by using elevated (25 mM) Mg2+ levels in the assay, or when Mg2+ was replaced by 5 mM Mn2+ in the assay. In contrast with this, potentiation of Gi function led to an increase in the lag period, as seen under conditions of agonist occupancy of inhibitory alpha 2-adrenoceptors (increase approximately 74%) or with the addition of 100 mM NaCl to the assays (increase approximately 44%). The local anaesthetic and membrane-fluidizing agent benzyl alcohol elicited both a profound decrease (around 70% at 80 mM) in the p[NH]ppG-induced lag period and a marked augmentation (around 5-fold) in the steady-state adenylate cyclase activity. When adenylate cyclase assays were done at 35 degrees C instead of 25 degrees C, then the lag period for activation by p[NH]ppG was decreased by around 33% and the steady-state rate increased by around 3-fold. At 35 degrees C, the addition of benzyl alcohol led to the apparent abolition of the lag period for p[NH]ppG activation of adenylate cyclase and amplified the steady-state rate by only around 2.2-fold. It is shown that Gi plays a fundamental role in determining the rate of activation of Gs. The proposal is formulated that such an action may be mediated through the release of beta gamma-subunits. Thus beta gamma-subunit dissociation is proposed as providing the rate-limiting step in Gi activation.</jats:p> |
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| institution | DE-Gla1, DE-Zi4, DE-15, DE-Pl11, DE-Rs1, DE-105, DE-14, DE-Ch1, DE-L229, DE-D275, DE-Bn3, DE-Brt1, DE-Zwi2, DE-D161 |
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| mega_collection | Portland Press Ltd. (CrossRef) |
| physical | 945-949 |
| publishDate | 1993 |
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| publisher | Portland Press Ltd. |
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| series | Biochemical Journal |
| source_id | 49 |
| spelling | Spence, S Houslay, M D 0264-6021 1470-8728 Portland Press Ltd. Cell Biology Molecular Biology Biochemistry http://dx.doi.org/10.1042/bj2910945 <jats:p>The non-hydrolysable GTP analogue guanylyl 5′-imidodiphosphate (p[NH]ppG) elicited a profound increase in the adenylate cyclase activity of human platelets. This occurred after a well-defined lag period of around 6 min, whereupon an enhanced steady-state rate was evident. The duration of the lag period was unchanged over a range of concentrations of p[NH]ppG which gave very different steady-state rates of adenylate cyclase activity. Prior activation of the stimulatory G-protein Gs by cholera-toxin pre-treatment abolished the lag period and elicited a small increase in the steady-state rate. Manipulating function of the inhibitory G-protein Gi also led to profound changes in the lag periods. Thus marked decreases in the lag were seen (approximately 70-81%) when Gi function was ablated through pre-treatment of platelet membranes with pertussis toxin, or by using elevated (25 mM) Mg2+ levels in the assay, or when Mg2+ was replaced by 5 mM Mn2+ in the assay. In contrast with this, potentiation of Gi function led to an increase in the lag period, as seen under conditions of agonist occupancy of inhibitory alpha 2-adrenoceptors (increase approximately 74%) or with the addition of 100 mM NaCl to the assays (increase approximately 44%). The local anaesthetic and membrane-fluidizing agent benzyl alcohol elicited both a profound decrease (around 70% at 80 mM) in the p[NH]ppG-induced lag period and a marked augmentation (around 5-fold) in the steady-state adenylate cyclase activity. When adenylate cyclase assays were done at 35 degrees C instead of 25 degrees C, then the lag period for activation by p[NH]ppG was decreased by around 33% and the steady-state rate increased by around 3-fold. At 35 degrees C, the addition of benzyl alcohol led to the apparent abolition of the lag period for p[NH]ppG activation of adenylate cyclase and amplified the steady-state rate by only around 2.2-fold. It is shown that Gi plays a fundamental role in determining the rate of activation of Gs. The proposal is formulated that such an action may be mediated through the release of beta gamma-subunits. Thus beta gamma-subunit dissociation is proposed as providing the rate-limiting step in Gi activation.</jats:p> The role of Gi and the membrane-fluidizing agent benzyl alcohol in modulating the hysteretic activation of human platelet adenylate cyclase by guanylyl 5′-imidodiphosphate Biochemical Journal |
| spellingShingle | Spence, S, Houslay, M D, Biochemical Journal, The role of Gi and the membrane-fluidizing agent benzyl alcohol in modulating the hysteretic activation of human platelet adenylate cyclase by guanylyl 5′-imidodiphosphate, Cell Biology, Molecular Biology, Biochemistry |
| title | The role of Gi and the membrane-fluidizing agent benzyl alcohol in modulating the hysteretic activation of human platelet adenylate cyclase by guanylyl 5′-imidodiphosphate |
| title_full | The role of Gi and the membrane-fluidizing agent benzyl alcohol in modulating the hysteretic activation of human platelet adenylate cyclase by guanylyl 5′-imidodiphosphate |
| title_fullStr | The role of Gi and the membrane-fluidizing agent benzyl alcohol in modulating the hysteretic activation of human platelet adenylate cyclase by guanylyl 5′-imidodiphosphate |
| title_full_unstemmed | The role of Gi and the membrane-fluidizing agent benzyl alcohol in modulating the hysteretic activation of human platelet adenylate cyclase by guanylyl 5′-imidodiphosphate |
| title_short | The role of Gi and the membrane-fluidizing agent benzyl alcohol in modulating the hysteretic activation of human platelet adenylate cyclase by guanylyl 5′-imidodiphosphate |
| title_sort | the role of gi and the membrane-fluidizing agent benzyl alcohol in modulating the hysteretic activation of human platelet adenylate cyclase by guanylyl 5′-imidodiphosphate |
| title_unstemmed | The role of Gi and the membrane-fluidizing agent benzyl alcohol in modulating the hysteretic activation of human platelet adenylate cyclase by guanylyl 5′-imidodiphosphate |
| topic | Cell Biology, Molecular Biology, Biochemistry |
| url | http://dx.doi.org/10.1042/bj2910945 |