File S1 - The Dimerization Domain in DapE Enzymes Is required for Catalysis

A. Analysis of the oligomeric state by size exclusion chromatograpy. Chromatogram showing elution of three DapE proteins (WT-HiDapE (blue), HiDapET (red), VcDapET(green)) from the calibrated column. The inset shows the calibration curve obtained by plotting Kav versus logMW forthe following standard proteins: Aprotinin (6.5 kDa, I), ribonuclease A (13.7 kDa, II), carbonic anhydrase (29 kDa, III) ovalbumin (43 kDa, IV), conalbumin (75 kDa,V), aldolase (158 kDa, VI), ferritin (440 kDa, VII), and thyroglobulin (669 kDa, VIII). B. The secondary structure of WT DapE and T325 mutants were measured using OlisDSM 20 circular dichrometer. Data was collected at room temperature with constant nitrogenflow every 1 nm, in the wavelength range of 190–260 nm. All samples were measured in acylindrical quartz cuvette with a 1 mm pathlength in 10 mM phosphate buffer pH 7.0. Threerepetitive scans were averaged, smoothed and background-subtracted for each measurement. Millidegree values obtained were converted to molar ellipticity (deg·cm2·dmol-1) by using the equation: Molar ellipticity = moM/10×L×C, where mo is millidegrees, M is molecular weight (g/mol), L is path length of cuvette (cm) and C is concentration (g/L).
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