TABLE 2.

Kinetic parameters for non-mutated (WT) and mutated recombinant NeuTTM

Data are the mean ± S.D.; n = 3–4.

Enzyme prepraration	Substrate	Conditions of experiment	Kmapp	Vmax	kcat	Kcat/Km
			μm	μmol min−1mg−1	s−1	s−1/M−1
WT	PPPi	50 °C, pH 9.7	40 ± 15	910 ± 70	288	7.2 106
His-tagged		Mg2+ = 5 mm
		37 °C, pH 9.7	21 ± 3	240 ± 30	76	3.6 106
		Mg2+ = 5 mm
		37 °C, pH 7.1	58 ± 5	60 ± 2	19	0.33 106
		Mg2+ = 5 mm
	ATP	50 °C, pH 8.1	800 ± 120	1.2 ± 0.2	0.36	0.45 103
		Mn2+ = 10 mm
WT untagged	PPPi	50 °C, pH 9.7m Mg2+ = 5 mm	100 ± 20	25,000 ± 2,000	7900	79 106
K8A	PPPi	50 °C, pH 9.7	390 ± 30	2,800 ± 500	887	2.3 106
His-tagged		Mg2+ = 5 mm
	PPPi	37 °C, pH 9.7	280 ± 55	235 ± 30	74	0.19 106
		Mg2+ = 5 mm
	ATP	50 °C, pH 10.1	1200 ± 500	12.5 ± 3.2	3.96	3.3 103
		Mn2+ = 10 mm
K85A	PPPi	50 °C, pH 9.7	720 ± 60	65 ± 7	21	29 103
His-tagged		Mg2+ = 5 mm
	PPPi	37 °C, pH 9.7	2600 ± 400	33 ± 3	10	4 103
		Mg2+ = 5 mm
K52R	PPPi	37 °C, pH 9.7	191 ± 8	3.1 ± 0.1	0.98	5.1 103
His-tagged		Mg2+ = 5 mm