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. 2003 Sep;185(18):5527–5535. doi: 10.1128/JB.185.18.5527-5535.2003

TABLE 2.

ATPase activities of MB and soluble F1Cp or F1FoCp after extraction with EDTA and detergenta

Treatment Protein (mg) Activityb
Activity (U · mg−1) in the presence of c:
U U · mg−1 DCCD (100 μM) Na azide (1 mM)
None 9.40 0.800 0.085 0.045 0.024
Membrane + DM
    Residual membrane 4.39 0.108 0.024 0.020 0.019
    Extracts (F1Fo) 5.24 1.198 0.228 0.125 0.073
Membrane + EDTA
    Residual membrane 7.32 0.240 0.033 0.022 0.020
    Extracts (F1) 2.22 0.942 0.424 0.330 0.135
a

For solubilization of F1Fo, washed membranes (2 mg/ml) were suspended in TMG buffer containing Na ATP (1 mM), DM (1% [wt/vol]), and PMSF (0.1 mM); the suspension was incubated on ice for 1 h and centrifuged at 100,000 × g for 1 h. The supernatant containing soluble F1Fo was collected, and the pellet containing residual membranes was washed and resuspended in TMG buffer. For solubilization of F1, washed membranes (1 mg/ml) were subjected to similar treatments, except that the membranes were suspended in 100 mM Tris-HCl (pH 8.0)-15 mM EDTA-2 mM LiCl-1 mM ATP. EDTA was removed from the extracts by dialysis against TMG buffer containing 1 mM Na ATP. The pellet containing residual membranes was washed and resuspended in TMG buffer. ATPase assays were done with Na ATP (2 mM) as a substrate.

b

One unit of activity is defined as 1 μmol of P1 released per min.

c

DCCD or Na azide was added to assay mixtures, which were incubated at 37°C for 10 min prior to the start of ATPase assays by the addition of ATP.