Inhibition of Na⁺/K⁺-ATPase and Mg²⁺-ATPase by metal ions and prevention and recovery of inhibited activities by chelators

Figures & data

Table I.  Experimental and recalculated “free” IC₅₀ values (μM).

	IC50 values (μM)
	Na^+/K^+-ATPase		Mg^2+-ATPase
Metal ion	Exp.	Calc.	Exp.	Calc.
Fe^2+	34	–	170	–
Co^2+^a	168	75	262	136
Cu^2+^b	7.1	0.6	42	33
Zn^2+	22	13	108	53
Hg^2+	0.7	–	3.6	–
Pb^2+	7.5	–	13.6	–

IC₅₀ values; ^aref. 5; ^bref. 6.

Figure 1 The dependence of MgATP^2– concentration in the reaction mixture containing 2 mM ATP and 5 mM MgCl₂ on the experimental Me²⁺ concentration: (▾) Cu²⁺; (▴) Co²⁺; (▪) Zn²⁺; (•) Fe²⁺, Hg²⁺, Pb²⁺.

Table II.  Inhibition of Na⁺/K⁺-ATPase and Mg²⁺-ATPase activity by simultaneous exposure to metal ions in mixtures.

	% Inhibition
	Na^+/ K^+-ATPase		Mg^2+-ATPase
Concentration (μM)	Experimental	Calculated	Experimental	Calculated
0.5 Cu +2 Fe	16.6 ± 2.5	17.2 ± 1.6	7.2 ± 1.3	5.2 ± 1.6
0.5 Cu +20 Fe	26.3 ± 2.8	28.6 ± 3.2	22.5 ± 2.1	3.2 ± 0.8
1 Cu +2 Fe	18.9 ± 1.1	20.4 ± 1.2	7.3 ± 0.9	5.0 ± 0.5
1 Cu +20 Fe	30.7 ± 2.2	32.8 ± 1.3	22.5 ± 2.9	6.4 ± 1.7
0.5 Cu +2 Zn	41.2 ± 2.5	11.6 ± 3.1	12.4 ± 2.4	4.1 ± 1.1
0.5Cu +20 Zn	65.9 ± 4.2	46.2 ± 2.1	17.8 ± 2.4	4.3 ± 0.9
1 Cu +2 Zn	68.4 ± 5.1	13.3 ± 2.3	13.9 ± 3.6	5.2 ± 1.5
1 Cu +20 Zn	100 ± 6.5	49.9 ± 3.6	18.7 ± 1.9	5.5 ± 1.0
2 Fe +2 Zn	20.8 ± 2.2	10.8 ± 2.1	1.2 ± 0.3	3.0 ± 0.2
2 Fe +20 Zn	85.1 ± 3.6	50.4 ± 1.1	3.9 ± 0.7	7.3 ± 1.3
20 Fe +2 Zn	65.1 ± 1.3	40.3 ± 1.4	1.6 ± 0.4	21.1 ± 2.1
20 Fe +20 Zn	100 ± 2.1	61.9 ± 3.3	3.6 ± 0.5	22.3 ± 1.9
1 Pb +0.5 Hg	42.2 ± 2.0	44.5 ± 1.5	14.2 ± 0.3	23.0 ± 1.3
5 Pb +0.5 Hg	75.5 ± 3.3	73.1 ± 1.2	38.7 ± 0.7	53.1 ± 3.3

Figure 2 Inhibition of Na⁺/K⁺-ATPase by Zn²⁺ in the absence (○) and presence of: 10 mM l-Cysteine (▪); 10 mM glutathione (▴) and 1 mM EDTA (•).

Figure 3 Inhibition of Na⁺/K⁺-ATPase by Hg²⁺ in the absence (○) and presence of: 1 mM EDTA (•); 10 mM l-cysteine (▪) and 10 mM glutathione (▴).

Figure 4 Effects of chelators: EDTA (•); l-cysteine (▪) and glutathione (▴) on the recovery of Na⁺/K⁺-ATPase activity in the presence of 1 × 10^–4 M ZnCl₂.

Table III.  Kinetic analysis of Na⁺/K⁺-ATPase and Mg²⁺-ATPase activity in the absence (control) and presence of metal ions.

			Mg^2+-ATPase
	Na^+/K^+-ATPase		High affinity		Low affinity
	Km (mM)	Vmax (μM Pi/mg/h)	Km (mM)	Vmax (μMPi/mg/h)	S0.5	Vmax (μM Pi/mg/h)
Control	0.69 ± 0.05	46.10 ± 2.20	0.32 ± 0.05	68.18 ± 1.20	3.79 ± 0.06	19.80 ± 0.90
Fe^2+	0.72 ± 0.09	25.80 ± 0.03	0.32 ± 0.04	57.80 ± 0.05	3.70 ± 0.04	19.80 ± 0.20
Co^2+	0.75 ± 0.09	33.20 ± 0.12	0.31 ± 0.03	55.20 ± 0.09	3.72 ± 0.06	18.90 ± 0.90
Cu^2+	0.74 ± 0.05	31.80 ± 0.80	0.33 ± 0.02	54.00 ± 1.20	3.64 ± 0.10	15.60 ± 1.00
Zn^2+	0.70 ± 0.09	19.60 ± 0.20	0.33 ± 0.01	56.70 ± 0.30	3.73 ± 0.03	19.73 ± 0.90

Experimental concentration of metal ions: Zn²⁺ = 1.5 × 10^–5 M; Cu²⁺ = 7.5 × 10^–6 M; Fe²⁺ = 3.4 × 10^–5 M; Co²⁺ = 1 × 10^–4 M.

Figure 5 (a) The dependence of Mg²⁺-ATPase activity on MgATP^2– in the absence (▴) and in the presence (Δ) of 15 μM ZnCl₂. Symbols represent experimental points. (b) The Mg²⁺-ATPase theoretical kinetic curves (Mg²⁺-ATPase activity vs MgATP^{2 −} concentration) of: “high affinity “Mg²⁺-ATPase subtype (○) in the absence of ZnCl₂ (•) and in the presence of ZnCl₂ (○); and “low affinity” Mg²⁺-ATPase subtype (squares) in the absence of ZnCl₂ (▪) and in the presence of ZnCl₂ (▪ or □).