Table 1 Kinetic parameters and catalytic efficiency of PfISN1 wild-type and mutantsa.

From: Structure and catalytic regulation of Plasmodium falciparum IMP specific nucleotidase

Protein

Substrate (modulator)

pH

kcat(app) (s−1)

Km(app) (mM)

(kcat/Km)(app) (mM−1 s−1)

Wild-type

IMP

5

10.3 ± 1.9

0.34 ± 0.09

29.8 ± 9.6

8

13.5 ± 0.8

66.0 ± 6.3

0.21 ± 0.02

AMP

5

23.2 ± 1.5

105.5 ± 13.4

0.22 ± 0.03

8

12.5 ± 0.9

72.7 ± 2.6b

0.17 ± 0.01

pNPP

8

0.0026 ± 0.0001

5.9 ± 0.5b

0.00045 ± 0.000042

IMP (ATP)c

8

21.0 ± 1.1

6.8 ± 1.1

3.1 ± 0.5

AMP (ATP)c

8

19.4 ± 0.6

71.7 ± 2.8b

0.27 ± 0.01

pNPP (ATP)c

8

0.0037 ± 0.0005

5.3 ± 0.7b

0.0007 ± 0.00013

ΔN30

IMP

5

54.8 ± 5.7

2.8 ± 0.7

19.6 ± 5.3

8

28.3 ± 0.3

29.9 ± 1.8

0.95 ± 0.06

ΔC10

5

55.1 ± 2.5

1.6 ± 0.3

34.4 ± 6.6

8

35.5 ± 1.2

20.7 ± 0.6

1.71 ± 0.08

K41L

5

22.5 ± 4.4

3.2 ± 1.3

7.0 ± 3.2

8

14.5 ± 0.8

15.6 ± 2.8

0.93 ± 0.17

H398V

5

32.5 ± 3.4

1.6 ± 0.6

20.3 ± 7.9

8

16.5 ± 0.8

10.9 ± 1.0

1.5 ± 0.2

D172N

pNPP

8

0.28 ± 0.01

1.51 ± 0.15

0.19 ± 0.02

D172A

8

0.019 ± 0.003

4.8 ± 1.4

0.0039 ± 0.0013

  1. aValues represent mean and SD of the fitted parameters obtained from three independent measurements.
  2. The plots from which the parameters in this table were derived are shown in Supplementary Figs. 6 and 7. The assays at pH 8.0 were carried out in 50 mM Tris-HCl, 30 mM MgCl2, at pH 5.0 in 50 mM MES, 30 mM MgCl2. The reaction mixture containing the substrate was mixed with 0.4 µM enzyme and incubated for 5 min at 25 °C. The liberated phosphate was estimated as described in “Methods”. Reactions with pNPP as substrate were continuously monitored at 405 nm.
  3. bThe values are K0.5.
  4. c4 mM ATP.
Back to article page