TABLE 1.

Purification and ATPase activity of wild type MDR3, the EQ/EQ, and the Q1174E mutant

Purified protein	Yield	No. of purifications	Km (MgATP)	vmax	kcat
	mg/100 g wet cell weight		mm	nmol min−1 mg−1	s−1
Wild type	6.3 ± 1.2	8	2.17 ± 0.20	354 ± 13	0.83 ± 0.03
			1.78 ± 0.10a	536 ± 11a	1.26 ± 0.03a
Wild type-BODIPY			1.26 ± 0.10	186 ± 6	0.44 ± 0.01
			1.43 ± 0.19a	175 ± 10a	0.41 ± 0.02a
E558Q/E1207Q	3.4 ± 0.6	5	b	b	b
Q1174E	2.0 ± 0.2	5	1.04 ± 0.15	286 ± 16	0.64 ± 0.04
Q1174E-BODIPY			0.74 ± 0.10	198 ± 5	0.46 ± 0.01

^a ATPase activity was in the presence of 300 μm DOPC lipids. The subtraction of the ATPase activity in the absence and presence of DOPC lipids revealed a Δv_max of 182 nmol min⁻¹ mg⁻¹ and Δk_cat of 0.43 s⁻¹ for wild type, a Δv_max of −9 nmol min⁻¹ mg⁻¹, and a Δk_cat of ≤ 0.03 s⁻¹ for labeled wild type MDR3-BODIPY.

^b The observed ATPase activity of the EQ/EQ mutant results from co-purified, contaminating ATPase. Please see text for further details.