Extended Data Fig. 5: Transport of phenoxyacetic acids and identification of critical residues for Substrate interaction.
From: Transport of phenoxyacetic acid herbicides by PIN-FORMED auxin transporters
a) Normalized representative current traces elicited by 1mM of the substrates. The perfusion protocol is shown on the top. b) Raw current traces of three representative solution exchange experiments used for KD determination of 2,4-D. For clarity the graph only shows three of the concentrations used to obtain the KD. The perfusion protocol is shown on the top. c) Relative transport rates of [3H]-4-CPA export by PIN3 from oocytes in the presence (▼) or absence (•) of PID. Initial internal 4-CPA concentration was 1 μM. Data points are mean ± SE transport rates using oocytes collected from different females. Groups were compared by a two-sided unpaired Student’s t-test (PIN3 versus PIN3 + PID, * P = 0.0.4101. d) Full width half maximum (FWHM) from the Peak current response determined by SSM electrophysiology on the wild-type PIN8 proteoliposomes of LPR10 (•) and LPR50 (▲) derived from 2mM of indicated substrate. Data are mean ± s.e.m. of n = 3 individual sensors. Groups were compared by a two-sided unpaired Student’s t-test (IAALPR10 versus IAALPR50: **** P < 0.0001; 2,4-DLPR10 versus 2,4-DLPR50: * P = 0.0148; 4-CPALPR10 versus 4-CPALPR50: *** P = 0.0003; 2-CPALPR10 versus 2-CPALPR50: * P = 0.0127; NPALPR10 versus NPALPR50: ** P = 0.0032). e) Relative Peak current response compared to WT elicited by 100μM (•) or 2mM (■) of indicated substrates using SSM electrophysiology on PIN8 N117A proteoliposomes. Data are mean ± s.e.m. of n = 3 individual sensors. There are no significant differences between the substrates at both concentrations used (One way ANOVA p = 0.3). Letters indicate the same significance level. f) Peak current response as a function of [2-CPA] determined by SSM electrophysiology on PIN8 N223A proteoliposomes. A Michaelis–Menten model is fit to describe kinetics (\({r}_{2-{CPA}}^{2}\) = 0.98; \({{EC}}_{50}^{2-{CPA}}\) 263 ± 64 μM; \({I}_{\max }^{2-{CPA}}\) 3 ± 0.1 nA; data points represent mean ± s.e.m.; n = 3 individual sensors).
