Figure 2.

Measurements for BetP in E. coli polar lipid proteoliposomes with spin labels introduced at positions 450 and 516. (A) Uptake of betaine in nanomolar per milligram protein was measured at 0.6 osmol/kg as a function of time for WT and MTSSL-labeled BetP cysteine variant C252T/G450C/S516C reconstituted into E. coli polar lipid liposomes. Uptake was initiated by adding saturating concentrations of 50 µM [¹⁴C]-betaine. Each value is the mean ± SEM of at least six independent measurements. (B) PELDOR normalized echo amplitude (Norm. Echo Amp.) time traces and assumed background traces (thinner smooth lines) for MTSSL-labeled BetP measured in the presence of 500 mM NaCl (black) or 300 mM NaCl and 5 mM betaine (green). (C and D) Probability of a distance P(r) versus distance (r) between spins derived from Tikonov regularization of the PELDOR time traces in B, before (C) and after (D) subtraction of a Gaussian distribution with a peak at ∼60 Å (see Fig. S2).