. Author manuscript; available in PMC: 2018 Jun 20.

Published in final edited form as: Biochemistry. 2017 Jun 7;56(24):3058–3067. doi: 10.1021/acs.biochem.7b00334

Table 1.

Equilibrium Titration and Kinetic Parameters for the Ligand Binding to WT, Opt-WT, and Cys-less CYP3A4

	WT	opt-WT	Cys-less
Equilibrium Titrations, K_s (μM)
RIT	0.05^a	0.045 ± 0.008	0.047 ± 0.012
BEC	0.3^b	0.77 ± 0.14	0.45 ± 0.06
MDZ
0.1 M phosphate	4.8^c	3.6 ± 0.1	0.73 ± 0.09
0.6 M phosphate	1.8^c	1.6 ± 0.1	0.22 ± 0.04
Ligand Binding Kinetics
RIT
k_fast(s⁻¹)	1.24 ± 0.05 (70%)^d	0.68 ± 0.02 (33%)	0.31 ± 0.03 (33%)
k_slow(s⁻¹)	0.26 ± 0.02 (30%)	0.033 ± 0.003 (67%)	0.037 ± 0.005 (67%)
BEC
k_fast(s⁻¹)	0.17 ± 0.02 (52%)	0.016 ± 0.003 (49%)	0.007 ± 0.001 (54%)
k_slow(s⁻¹)	0.0089 ± 0.0014 (48%)	0.0016 ± 0.0004 (51%)	0.0013 ± 0.003 (46%)
MDZ, 0.1 M phosphate
k_fast(s⁻¹)	1.50 ± 0.05 (70%)	0.19 ± 0.03 (43%)	0.15 ± 0.02 (40%)
k_slow(s⁻¹)	0.022 ± 0.004 (30%)	0.021 ± 0.003 (57%)	0.022 ± 0.03 (60%)
MDZ, 0.6 M phosphate
k_fast(s⁻¹)	3.4 ± 0.3 (82%)	0.26 ± 0.03 (49%)	0.27 ± 0.02 (43%)
k_slow(s⁻¹)	0.23 ± 0.04 (18%)	0.027 ± 0.004 (51%)	0.036 ± 0.003 (57%)
First Electron Transfer from CPR
k_fast(min⁻¹)	1.20 ± 0.05 (65%)	0.72 ± 0.02 (85%)	1.22 ± 0.03 (64%)
k_slow(min⁻¹)	0.26 ± 0.02 (35%)	0.030 ± 0.003 (15%)	0.22 ± 0.02 (36%)

Determined previously.³

Determined previously.⁴

Determined previously.²⁴

Percentage of absorbance change observed during the indicated kinetic phase.