FIGURE 3.

Activation of LpxE by solubilization of Novablue(DE3)/pLpxE-4 membranes with 4% Triton X-100. Membranes of Novablue(DE3)/pLpxE-4 were solubilized with 4% Triton X-100, ultracentrifuged to remove insoluble material, and assayed for 1-phosphatase activity with Kdo₂-[4′-³²P]lipid IV_A as substrate under otherwise standard conditions at 30 °C. The no enzyme control is shown in the 1st lane. Membranes from Novablue (DE3)/pLpxE-4 that had not been solubilized with Triton X-100 were used at 0.5 mg/ml in the 2nd and 3rd lanes. Triton X-100-solubilized supernatant of Novablue(DE3)/pLpxE-4 membranes was used at 0.5 mg/ml in the 4th and 5th lanes. At each time point, a 4-μl portion was quenched, and the extent of dephosphorylation was assessed by TLC and PhosphorImager analysis.