Abstract
Purified bovine renal papillary Na+ channels, when reconstituted into planar lipid bilayers, reside in three conductance states: a 40-pS main state, and two subconductive states (12-13 pS and 24-26 pS). The activity of these channels is regulated by phosphorylation and by G-proteins. Protein kinase A (PKA)-induced phosphorylation increased channel activity by increasing the open state time constants from 160 ± 30 (main conductance), and 15 ± 5 ms (both lower conductances), respectively, to 365 ± 30 ms for all of them, PKA phosphorylation also altered the closed time of the channel from 250 ± 30 ms to 200 ± 35 ms, thus shifting the channel into a lower- conductance, long open time mode. PKA phosphorylation increased the P(Na):P(K) of the channel from 7:1 to 20:1, and shifted the amiloride inhibition curve to the right (apparent K(i)/(amil) from 0.7 to 20 μM). Pertussis toxin-induced ADP-ribosylation of either phosphorylated or nonphosphorylated channels decreased the P(Na):P(K) to 2:1 and 4:1, respectively, and altered K(i)/(amil) to 8 and 2 μM for phosphorylated and nonphosphorylated channels, respectively. GTP-γ-S treatment of either phosphorylated or nonphosphorylated channels resulted in an increase of P(Na):P(K) to 30:1 and 10:1, respectively, and produced a leftward shift in the amiloride dose-response curve, altering K(i)/(amil) to 0.5 and 0.1 μM, respectively. These results suggest that amiloride-sensitive renal Na+ channel biophysical characteristics are not static, but depend upon the biochemical state of the channel protein and/or its associated G-protein.
| Original language | English |
|---|---|
| Pages (from-to) | 1789-1800 |
| Number of pages | 12 |
| Journal | Biophysical Journal |
| Volume | 69 |
| Issue number | 5 |
| Publication status | Published - 1995 |
| Externally published | Yes |
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ASJC Scopus subject areas
- Biophysics
Cite this
Biochemical status of renal epithelial Na+ channels determines apparent channel conductance, ion selectivity, and amiloride sensitivity. / Ismailov, I. I.; Berdiev, B. K.; Benos, D. J.
In: Biophysical Journal, Vol. 69, No. 5, 1995, p. 1789-1800.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Biochemical status of renal epithelial Na+ channels determines apparent channel conductance, ion selectivity, and amiloride sensitivity
AU - Ismailov, I. I.
AU - Berdiev, B. K.
AU - Benos, D. J.
PY - 1995
Y1 - 1995
N2 - Purified bovine renal papillary Na+ channels, when reconstituted into planar lipid bilayers, reside in three conductance states: a 40-pS main state, and two subconductive states (12-13 pS and 24-26 pS). The activity of these channels is regulated by phosphorylation and by G-proteins. Protein kinase A (PKA)-induced phosphorylation increased channel activity by increasing the open state time constants from 160 ± 30 (main conductance), and 15 ± 5 ms (both lower conductances), respectively, to 365 ± 30 ms for all of them, PKA phosphorylation also altered the closed time of the channel from 250 ± 30 ms to 200 ± 35 ms, thus shifting the channel into a lower- conductance, long open time mode. PKA phosphorylation increased the P(Na):P(K) of the channel from 7:1 to 20:1, and shifted the amiloride inhibition curve to the right (apparent K(i)/(amil) from 0.7 to 20 μM). Pertussis toxin-induced ADP-ribosylation of either phosphorylated or nonphosphorylated channels decreased the P(Na):P(K) to 2:1 and 4:1, respectively, and altered K(i)/(amil) to 8 and 2 μM for phosphorylated and nonphosphorylated channels, respectively. GTP-γ-S treatment of either phosphorylated or nonphosphorylated channels resulted in an increase of P(Na):P(K) to 30:1 and 10:1, respectively, and produced a leftward shift in the amiloride dose-response curve, altering K(i)/(amil) to 0.5 and 0.1 μM, respectively. These results suggest that amiloride-sensitive renal Na+ channel biophysical characteristics are not static, but depend upon the biochemical state of the channel protein and/or its associated G-protein.
AB - Purified bovine renal papillary Na+ channels, when reconstituted into planar lipid bilayers, reside in three conductance states: a 40-pS main state, and two subconductive states (12-13 pS and 24-26 pS). The activity of these channels is regulated by phosphorylation and by G-proteins. Protein kinase A (PKA)-induced phosphorylation increased channel activity by increasing the open state time constants from 160 ± 30 (main conductance), and 15 ± 5 ms (both lower conductances), respectively, to 365 ± 30 ms for all of them, PKA phosphorylation also altered the closed time of the channel from 250 ± 30 ms to 200 ± 35 ms, thus shifting the channel into a lower- conductance, long open time mode. PKA phosphorylation increased the P(Na):P(K) of the channel from 7:1 to 20:1, and shifted the amiloride inhibition curve to the right (apparent K(i)/(amil) from 0.7 to 20 μM). Pertussis toxin-induced ADP-ribosylation of either phosphorylated or nonphosphorylated channels decreased the P(Na):P(K) to 2:1 and 4:1, respectively, and altered K(i)/(amil) to 8 and 2 μM for phosphorylated and nonphosphorylated channels, respectively. GTP-γ-S treatment of either phosphorylated or nonphosphorylated channels resulted in an increase of P(Na):P(K) to 30:1 and 10:1, respectively, and produced a leftward shift in the amiloride dose-response curve, altering K(i)/(amil) to 0.5 and 0.1 μM, respectively. These results suggest that amiloride-sensitive renal Na+ channel biophysical characteristics are not static, but depend upon the biochemical state of the channel protein and/or its associated G-protein.
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M3 - Article
VL - 69
SP - 1789
EP - 1800
JO - Biophysical Journal
JF - Biophysical Journal
SN - 0006-3495
IS - 5
ER -