Table 2.

Effect of mutations of Arg275 on the substrate specificity of hCPO

	Substrate
Mutation	FA-EE	FA-AK	FA-FF
hCPOΔC WT
KM	374.10 ± 48.36	NM	NM
Vmax	0.23 ± 0.01	NM	NM
Kcat	7.86 ± 0.40	NM	NM
Kcat/KM	0.021 ± 0.004	NM	NM
hCPOΔC R275D
KM	NM	671.50 ± 112.20	NM
Vmax	NM	0.81 ± 0.07	NM
Kcat	NM	27.77 ± 2.39	NM
Kcat/KM	NM	0.041 ± 0.010	NM
hCPOΔC R275A
KM	NM	NM	357.80 ± 40.34
Vmax	NM	NM	0.41 ± 0.02
Kcat	NM	NM	14.15 ± 0.64
Kcat/KM	NM	NM	0.039 ± 0.006
CPB
KM	NM	400.00 ± 86.98	NM
Vmax	NM	0.70 ± 0.006	NM
Kcat	NM	23.17 ± 2.05	NM
Kcat/KM	NM	0.058 ± 0.018	NM
CPA
KM	NM	NM	136.60 ± 15.94
Vmax	NM	NM	0.44 ± 0.02
Kcat	NM	NM	87.65 ± 4.04
Kcat/KM	NM	NM	0.64 ± 0.104

Enzymatic activity measurements were performed in triplicate. Data are shown as mean ± SE. See SI Appendix, SI Materials and Methods for more details about the equations used for kinetic constant determinations. Units are: K_M: µM; V_max: µM·s⁻¹; K_cat: s⁻¹; and K_cat/K_M: µM⁻¹·s⁻¹. NM, not measurable levels of activity.