Table 1. Pyocyanin complementation of a P. aeruginosa PA14 pqsE− strain transformed with the PqsE variants and the predicted free energy variation caused by each point mutation.
PqsE Variant | Pyocyanin Production (mg/L) a | ΔΔG 0 (kcal/mol)b | SA | |
WT | 6.2 (0.2) | – | – | |
Active Site [AS] | H69A | 0.1 (0.1) | 2.9 | 0.0 |
H71A | 2.7 (0.6) | 1.5 | 0.2 | |
D73A | 4.1 (0.8) | 0.9 | 0.1 | |
H74A | 3.5 (0.6) | 2.3 | 0.0 | |
H159A | 0.1 (0.0) | 2.3 | 0.0 | |
D178A | 0.1 (0.1) | 1.4 | 0.0 | |
E182A | 1.2 (0.4) | 1.5 | 0.1 | |
L193A | 4.7 (1.2) | 2.3 | 0.1 | |
F195A | 0.8 (0.3) | 2.7 | 0.1 | |
H221A | 1.1 (0.2) | 2.3 | 0.1 | |
Q272A | 1.4 (0.4) | 0.9 | 0.2 | |
S273A | 0.1 (0.1) | 0.9 | 0.0 | |
F276A | 0.1 (0.0) | 2.3 | 0.0 | |
L277A | 4.5 (1.1) | 1.8 | 0.1 | |
S285A | 5.0 (0.6) | 0.3 | 0.1 | |
S285W | 5.6 (0.6) | 0.1 | 0.1 | |
R288A | 5.2 (1.0) | 0.5 | 0.5 | |
C-terminal α-helix motif [α8/α9] | L248A | 4.1 (1.1) | 2.2 | 0.0 |
L261A | 0.2 (0.1) | 2.0 | 0.0 | |
W269A | 0.1 (0.0) | 3.1 | 0.1 | |
H282A | 0.4 (0.2) | 2.0 | 0.1 | |
M286A | 0.1 (0.1) | 1.9 | 0.0 | |
L290A | 0.1 (0.1) | 2.5 | 0.0 | |
KH-like motif [KH] | R95A | 5.1 (0.5) | 0.6 | 0.5 |
K101A | 4.4 (0.4) | 0.6 | 0.6 | |
R107A | 4.0 (0.8) | 0.1 | 0.6 | |
R111A | 0.3 (0.2) | 0.2 | 0.6 | |
D130A | 0.2 (0.1) | 0.2 | 0.6 |
Pyocyanin production (mg/L) was measured in triplicate as described in the experimental procedures. The mean value (± standard deviation) is shown. b Free energy variations caused by point mutants of PqsE, as predicted by the PoPMuSiC software (v.2). c Calculated solvent accessibility for each WT residue, ranging from 0 (fully buried in the protein core) to 1 (fully accessible to the solvent).