Figure 4.
Influence of Ion Concentrations and pH on ATP Binding of I-2N.
α-32P-ATP binding was measured in a filter binding assay as described in Methods.
(A) to (C) The maximal amount of α-32P-ATP bound was set to 1.
(A) I-2N (1 μM) was incubated with 0.07 μM α-32P-ATP (6.3 × 105 cpm/pmol) in the standard reaction mixture except for a variable NaCl concentration.
(B) I-2N (1.5 μM) was incubated with 0.1 μM α-32P-ATP (4.3 × 105 cpm/pmol) in the standard reaction mixture except for a variable pH.
(C) I-2N (0.8 μM) was incubated with 0.2 μM α-32P-ATP (2.1 × 105 cpm/pmol) in the standard reaction mixture except for a variable MgCl2 concentration and the absence of DTT and EDTA.
(D) I-2N (0.8 μM) was incubated with 0.2 μM α-32P-ATP (2.1 × 105 cpm/pmol) in the standard reaction mixture except for variable concentrations of the divalent (Div.) cations of MgCl2, MnCl2, or CaCl2. The maximal amount of α-32P-ATP bound in presence of MgCl2 was set to 1.