Table 1.
Isolated AT1A mutants highly sensitive to CGP42112A
| Harbored amino acid substitutions | |||
|---|---|---|---|
| Major | Others |
EC50, nM | Basal IP, % |
| WT | 375 | 99 ± 2 | |
| F77S(2)† | 25.7 | 102 ± 1 | |
| F77S(2) | G203A-D273N | n.d. | n.d. |
| F77S(2) | (I38V)-W253R-(L316P) | n.d. | n.d. |
| F77Y(2) | W253R | 10.9 | 111 ± 1* |
| L78F(2) | L191P-L212H-F293L | 12.3 | 101 ± 4 |
| S107F(3) | 7.20 | 102 ± 1 | |
| F110C(3) | I201T-(F304I) | 11.3 | 90 ± 17 |
| N111S(3) | N231S-F293L | 7.18 | 170 ± 9* |
| L112H(3) | V29A-L143P | 15.0 | 141 ± 3* |
| L112H(3) | F44S-T175A-F182Y-K325* | 4.94 | 120 ± 1* |
| L112F(3) | V164A | 7.20 | 105 ± 2 |
| L118H(3) | (K318E) | 11.7 | 116 ± 1* |
| E173G(e2) | 21.1 | 92 ± 3 | |
| P162Q-A181V | 7.15 | 99 ± 4 | |
| I193K(5) | M142R-S186P | 11.2 | 104 ± 1 |
| L195P(5) | (G306R) | 7.94 | 118 ± 3* |
| L195R(5) | T178S-S329G | 14.8 | 100 ± 4 |
| T198I(5) | (K310N) | 10.1 | 99 ± 6 |
| I245T(6)† | 20.7 | 111 ± 1* | |
| I245T(6) | M142T-H256F | 4.36 | 149 ± 8* |
| I245T(6) | (F44L) | n.d. | n.d. |
| L305Q(7) | 7.39 | 143 ± 4* | |
| L305Q(7) | (S326T) | n.d. | n.d. |
| L305Q(7) | D9E | n.d. | n.d. |
| L305Q(7) | A221V | n.d. | n.d. |
| L305Q(7) | Q15R-V52D-S189P-Q229R | n.d. | n.d. |
Characteristics of the isolated clones (†, except for the F77S and I245T mutants, which were constructed subsequently): list of amino acid substitutions (one letter amino acid code; Z, stop codon), either resulting in much (“Major”) or slightly (underlined) higher sensitivity to CGP42112A, in no change (amino acid substitutions in brackets) or untested (others); EC50 of CGP42112A in the æquorin assay (representative of at least two independent determinations); basal IP production (see Fig. 2). The positions of the mutated amino acids (for major substitutions) in the putative secondary structure of AT1A receptor are indicated in brackets, either TM (2, 3, 5, 6, or 7) or second extracellular loop (e2).
Significantly different from WT.