Crystallization and data collection

C-terminal His-tagged EstDL136(ΔPMP) and EstDL136(ΔPMP/S156A) in buffer A were concentrated to 15 mg/ml. Crystallization of both enzymes was performed by the sitting-drop vapor-diffusion method at 295 K using a crystallization solution of 500 mM ammonium fluoride (pH 6.5), 30% PEG3350, 5% glycerol, and 120 mM TCEP. The binding of the Cm to EstDL136(ΔPMP/S156A) was achieved by soaking EstDL136(ΔPMP/S156A) crystals with 5 mM Cm (Sigma) in the crystallization solution for 50 min. Cryoprotection was performed using 20% (v/v) PEG400 for EstDL136(ΔPMP) and 3.5% (w/v) myo-inositol for EstDL136(ΔPMP/S156A).

X-ray diffraction data were collected at 100 K in the Pohang Accelerator Laboratory (Pohang, Korea). Data were processed using HKL2000 [16] and the details of the data collection are shown in Table 1.

^aNumbers in parentheses refer to data in the highest resolution shell.

^bThe CC_1/2 is the Pearson correlation coefficient (CC) calculated from each subset containing a random half of the measurements of unique reflection

^cR_work = Σ ||F_obs|-|F_cal||/ Σ|F_obs|

^dR_free is the same as R_obs for a selected subset (5%) of the reflections that was not included in prior refinement calculations.