FIGURE 5.

HPLC chromatograms of OatA reaction products. (A) Reactions containing ± OatA (80 μM), N-acetyl-serine (OAS, 1 mM), and AcCoA (1.2 mM) were incubated for 1 h at 37°C and OatA was removed from reaction mixture, which was then injected (100 μL) onto an Aminex HPX-87H column (Bio-Rad) as described under “Materials and Methods” section. Separation of desired product (DAS, retention time ∼14 min, red peak) was monitored using an absorbance of 215 nm. (B) Proper separation of DAS was compared to known standards: AcCoA (retention time between 5–7 min), CoA (retention time between 5 and 7 min), OAS (retention time 5–7 min), and NAS (retention time ∼12 min). All standards were re-suspended for a final concentration of 1 mM. Legends in (A) also apply to (B). Experiment was repeated in three independent experiments.