Table 2.

Amino Acid Substrate Specificity of wt TH and Selected Mutants

Protein (mutation)	Substrate	Vmax (nmol min−1mg−1)	Km (μM)
wt TH	l-Tyr	2460 ± 417	5.1 ± 2.2	484	88	114
	l-Phe	1640 ± 148	296 ± 69	5.5
	l-Dopa	126 ± 11	29.8 ± 8.5	4.2
p.Arg233His	l-Tyr	150 ± 30	13.0 ± 4.0	11.5	4.8	7.7
(c.698A>G)	l-Phe	352 ± 33	149 ± 22	2.4
	l-Dopa	1.6	1.0	1.5
p.Gly247Ser	l-Tyr	1189 ± 53	11.8 ± 0.9	101	16	36
(c.739G>A)	l-Phe	734 ± 46	119 ± 25	6.2
	l-Dopa	50 ± 9.4	17 ± 12	2.8
p.Cys359Phe	l-Tyr	329.4 ± 4.2	37.6 ± 1.1	8.8	80
(c.1076G>T)	l-Phe	73.2 ± 5.7	663 ± 99	0.1
	l-Dopa	n.m.	n.m.
p.Phe375Leu	l-Tyr	241 ± 20	56 ± 11	4.3	2.2	11
(c.1125C>G)	l-Phe	1960 ± 44	993 ± 37	2.0
	l-Dopa	77 ± 30	202 ± 135	0.4
p.Leu387Met	l-Tyr	2830 ± 1450	24 ± 17	118	21	131
(c.1159C>A)	l-Phe	1407 ± 78	258 ± 38	5.5
	l-Dopa	62 ± 14	68 ± 39	0.9
p.Gly414Arg	l-Tyr	66.8 ± 7.5	26.6 ± 8.1	2.5	0.5	1.3
(c.1240G>A)	l-Phe	452 ± 17	92 ± 12	4.9
	l-Dopa	35.1 ± 7.2	18 ± 13	1.9

The table summarizes the different enzyme kinetic parameters obtained for wt TH and some of the mutants, using l-Tyr, l-Phe, or l-Dopa as substrate. We did not observe substrate inhibition kinetics for l-Phe or l-Dopa, which is why only V_max and K_m values are reported. The data for l-Tyr are taken from Table1. Data points were fitted using standard MM kinetics by nonlinear regression (Sigma plot) as described in Materials and Methods section. Values are given as the best estimate ± standard error of estimate. n.m., not measured.