Journal of Cell and Molecular Biology - ResearchGate

pH and temperature studies
Temperature dependent studies showed that the
enzyme was optimally active at 40 o C (Figure 6).
Arrhenius plot of the log of initial velocity as a
function of the reciprocal of absolute temperature
gave an Ea of 6.27 kJ/mol (Figure 7). pH dependent
studies revealed that the enzyme was optimally
active at pH 5.0 (Figure 8).
Activity (µmol/min - )
0.16
0.14
0.12
0.1
0.08
0.06
0.04
0.02
0
0 20 40 60 80
Temperature 0 C
Figure 6. Optimum temperature determination for
the activity of Plasmodium berghei cysteine
protease.
Activity ( µmol/min)
1.4
1.2
1
0.8
0.6
0.4
0.2
0
0 5 10 15
pH
Figure 7. Optimum pH determination for the
activity of Plasmodium berghei cysteine protease.
1/V(µmol/
min)
60
50
40
30
20
10
y = 10.305x + 4.1076
0
-6 -4 -2 0 2 4
1/S(mg/ml)
6
Figure 8. Lineweaver-Burk plot relating
Plasmodium berghei Cysteine protease reaction
velocity to gelatin concentration. KM was calculated
as milligram gelatin /ml. Each point represents the
average of three experiments.
Characterization of Plasmodium berghei cysteine protease 47
Initial velocity studies
Lineweaver Burk plots of initial velocity studies of
the enzyme gave the KM and Vmax values of 2.5
mg/ml and 0.2 µmol/min, respectively (Figure 9).
Inhibitory studies conducted with the synthetic
compounds PP-56 and PP-54, currently being
validated for their anti-malarial activity, revealed
competitive inhibition patterns with Ki values of
48.88 µg/ml (Figure 10) and 0.14 µg/ml (Figure
11).
10
Log of activity
(µmol/min)
1
y = -0.1373x + 3.683
1 2 3 4 5 6
1/T (x10 -3 )
Figure 9. Arrhenius plot for the determination of
the Ea for Plasmodium berghei cysteine protease
activity.
1/V (µmol/min)
12
10
8
6
4
2
y = 4.9261x + 2.3458
y = 1.9315x + 2.1625
0
-1.5 -1 -0.5 0 0.5 1 1.5 2
1/S (mg/ml)
No Inhibitor Compd P56
Figure 10. Lineweaver-Burk plots of initial
velocity data for the determination of inhibition
pattern on Plasmodium berghei Cysteine protease
by compound PP-56 using gelatin as substrate. Data
from three experiments were used to plot the graph
using MS Excel program.