TABLE 3.
Biophysical parameters of CaV3.2 wild-type and double mutant channels were estimated from recordings obtained with the cut-open oocyte technique in the presence of 10 mm Ba2+
Double mutants are shown with the IIS4-S5 mutant always to the left of the pair. Most double mutants were made between IIS4-S5 and IIS6. Double mutants with Asn-1548 in IIIS6 and Asn-1853 in IVS6 are shown below and are separated by a line. Activation properties (E0.5,act and z) were estimated from the mean I-V relationships and fitted to a Boltzmann equation where z is the slope factor. Activation and inactivation energies were calculated as follows: ΔGact = z · F · E0.5,act; ΔΔGact = ΔGact,mut − ΔGact,wt; ΔGinact = z · F · E0.5,inact; and ΔΔGinact = ΔGinact,mut − ΔGinact,wt. The voltage dependence of inactivation was determined from the peak currents after 2-s depolarizing pulses from a holding potential of −120 mV. The data are shown with the mean ± S.E. of the individual experiments and the number of experiments appears in parentheses.
| V0.5, act | ΔGact | ΔΔGact | Ipeak | V0.5, inact | ΔGinact | τact, −20 mV | τinact, −20 mV | ΔΔGinter (act) | ΔΔGinteract | |
|---|---|---|---|---|---|---|---|---|---|---|
| mV | kcal mol−1 | nA | mV | kcal mol−1 | ms | kcal mol−1 | ||||
| CaV3.2 WT | −47 ± 1 (37) | −4.3 ± 0.2 | NAa | −1300 ± 200 | −65 ± 1 (10) | −8.6 ± 0.5 | 3.2 ± 0.1 (18) | 11.6 ± 0.2 | NA | NA |
| V906G/I1013G | −54 ± 2 (5) | −6.1 ± 0.7b | −1.8 ± 0.9 | −300 ± 100 | −69.1 ± 0.6 (4) | −7.9 ± 0.6 | 4.2 ± 0.3c (5) | 48 ± 2b | −0.8 ± 1.8 | −2.1 ± 2.5 |
| V907G/I1013G | −55 ± 1 (6) | −4.1 ± 0.2 | 0.2 ± 0.4 | −190 ± 40 | −78.2 ± 0.2 (3) | −7.8 ± 0.5 | 2.9 ± 0.3 (6) | 40 ± 1b | 4.6 ± 1.3 | 0.3 ± 2.1 |
| L908G/N1008G | −50 ± 3 (5) | −5.0 ± 0.7 | −0.7 ± 0.9 | −80 ± 20 | −76 ± 1 (4) | −6.9 ± 0.3 | 2.8 ± 0.3 (5) | 11 ± 1 | −1.3 ± 1.9 | −0.5 ± 1.9 |
| L908A/I1013A | −52 ± 1 (5) | −5.4 ± 0.3 | −1.1 ± 0.5 | −160 ± 40 | −67 ± 1 (5) | −5.8 ± 0.6c | 3.4 ± 0.3 (5) | 72 ± 2b | −1.3 ± 1.2 | 2.4 ± 2.5 |
| L908G/I1013G | −48 ± 1 (6) | −3.9 ± 0.3 | 0.4 ± 0.5 | −120 ± 20 | −72 ± 1 (4) | −7.2 ± 0.4 | 3.7 ± 0.5 (6) | 25.0 ± 0.8b | 2.1 ± 1.1 | 0.6 ± 2.5 |
| V909G/I1013G | −48 ± 1 (4) | −4.9 ± 0.9 | −0.6 ± 1.1 | −170 ± 40 | −65 ± 2 (3) | −6.9 ± 0.8 | 5.0 ± 0.5b (4) | 53 ± 3b | 0.4 ± 1.8 | 1.1 ± 2.6 |
| K910G/I1013G | −46.7 ± 0.6 (5) | −3.9 ± 0.2 | 0.4 ± 0.4 | −150 ± 30 | −65 ± 1 (4) | −6.2 ± 0.2 | 3.6 ± 0.2 (5) | 47 ± 4b | 0.2 ± 0.9 | 1.1 ± 2.3 |
| T911A/I1013A | −46 ± 2 (6) | −4.4 ± 0.4 | −0.1 ± 0.6 | −340 ± 80 | −60.5 ± 0.8 (6) | −6.9 ± 0.5 | 6.1 ± 0.5b (6) | 71 ± 5b | −1.6 ± 1.2 | NDd |
| T911G/I1013G | −43.4 ± 0.8 (6) | −3.6 ± 0.2 | 0.7 ± 0.4 | −260 ± 30 | −66 ± 1 (6) | −6.0 ± 0.5c | 6.7 ± 0.5b (6) | 41 ± 2b | 1.8 ± 0.9 | 2.9 ± 2.6 |
| T911G/L1014G | −47 ± 2 (6) | −3.8 ± 0.4 | 0.5 ± 0.6 | −200 ± 100 | −75 ± 1 (4) | −8 ± 1 | 8 ± 1b (6) | 26 ± 1b | −0.8 ± 1 | 1.8 ± 2.8 |
| M912G/I1013G | −48.6 ± 0.7 (6) | −4.7 ± 0.1 | −0.4 ± 0.3 | −180 ± 60 | −68 ± 1 (6) | −7.5 ± 0.4 | 5.0 ± 0.6b (6) | 42 ± 1b | 1.1 ± 1.1 | 2.3 ± 2.8 |
| N914G/E1016G | −36.5 ± 0.9 (6) | −2.8 ± 0.2b | 1.5 ± 0.4 | −90 ± 10 | −61.0 ± 0.7 (6) | −7.0 ± 0.4 | 3.9 ± 0.2c (6) | 15.0 ± 0.3b | −0.8 ± 1.2 | ND |
| V915G/L1010G | −27 ± 3 (6) | −1.6 ± 0.3b | 2.7 ± 0.5 | −110 ± 30 | −58 ± 1 (3) | −8.6 ± 0.6 | 5.1 ± 0.6b (6) | 14.9 ± 0.9b | −0.3 ± 0.6 | ND |
| V907A/N1548A | −43 ± 3 (4) | −3.4 ± 0.6 | 0.9 ± 0.8 | −50 ± 10 | −90 ± 2 (2) | −6 ± 2 | 3.1 ± 0.2 (4) | 12 ± 2 | 1 ± 1.2 | −1.4 ± 3.1 |
| V907G/N1548G | −47 ± 1 (10) | −3.8 ± 0.2 | 0.5 ± 0.4 | −110 ± 17 | −82 ± 2 (6) | −5.7 ± 0.6c | 2.6 ± 0.1 (10) | 15 ± 1b | 3.4 ± 1.2 | 1.6 ± 1.6 |
| L908A/N1548A | −39 ± 2 (6) | −3.6 ± 0.3 | 0.7 ± 0.5 | −180 ± 30 | −74.6 ± 0.9 (6) | −7.3 ± 0.2 | 4.0 ± 0.3c (6) | 9.9 ± 0.3c | 0.8 ± 1.0 | −0.8 ± 1.7 |
| L908G/N1548G | −35 ± 1 (11) | −3.3 ± 0.4c | 1.0 ± 0.6 | −220 ± 60 | −74 ± 1 (9) | −7.2 ± 0.5 | 3.5 ± 0.2 (11) | 16.5 ± 0.4b | 1.2 ± 1.1 | −0.3 ± 2.0 |
| T911A/N1548A | −36 ± 2 (7) | −2.9 ± 0.4b | 1.4 ± 0.6 | −240 ± 70 | −73 ± 2 (6) | −8.4 ± 0.5 | 4.0 ± 0.2c (7) | 10.0 ± 0.7c | 0.2 ± 1.0 | ND |
| T911G/N1548G | −37 ± 2 (7) | −2.6 ± 0.2b | 1.7 ± 0.4 | −100 ± 20 | −74 ± 2 (4) | −7.6 ± 0.4 | 3.4 ± 0.4 (7) | 13 ± 1 | 1.3 ± 0.8 | 0.4 ± 1.9 |
| L908G/N1853G | −43.4 ± 0.9 (9) | −3.8 ± 0.2 | 0.5 ± 0.4 | −180 ± 30 | −70 ± 2 (5) | −7.9 ± 0.6 | 3.5 ± 0.2 (9) | 17.2 ± 0.4b | −1 ± 1 | ND |
a NA, not applicable.
b p < 0.01 as compared with CaV3.2 WT.
c p < 0.001 as compared with CaV3.2 WT.
d ND, not determined.