H change at 0 °C when the buffer was in the system. Addition
of CO2 to the reaction mixture at time zero tended to drop the
pH of the mixture to the low values, but the presence of the
buffer balanced this effect, and the pH leveled off at a value
slightly lower than the pH of the buffered solution.
Since the initial rate of pH drop (rA) is equal to the slope of
the curves at the first few seconds of the reaction, to compare
the rate of pH drop at different concentrations, the slope of the
curves in Figure 1 were calculated for the early part of the
reaction and -1/slope, which is equivalent to 1/rA for each
enzyme concentration, as shown in Figure 2. It is clearly
observed in this figure that the pH drop depends on the
concentration of the enzyme. At higher enzyme concentrations,
pH drops more quickly. Also, the results show that in the
absence of the enzyme (control run) the rate of pH decrease is
much slower compared to the rate of pH drop in the presence
of the enzyme, even at low concentration of the enzyme (e.g.,
0.2 µM). These results demonstrate that enzyme carbonic
anhydrase increases the rate of hydration of CO2, even at low
concentrations.
The results of the experiments at 30 °C and in the presence
of buffer are shown in Figure 3. The same trend can be seen in
this figure, but in this case the rate of change in pH was higher
than that at 0 °C. The comparison of rates at different
concentrations at 30 °C is illustrated in Figure 4. These results
suggest that, at high temperatures, the concentration of the
enzyme is not as effective in the promotion of hydration of CO2
as it is in lower temperatures. This is beneficial because, in the
Table 1. Summary of Precipitation Experiments
set temp (°C) enzyme concn (µM) buffer wt of precipitate (g) no. of runs error (%)
1 0 3 yes 0.2098 2 0.1
2 0 6 yes 0.2106 4 4.2
3 0 6 no 0 3 N/A
4 0 no enzyme yes 0.19 2 1
5 30 3 yes 0.1280 2 0.16
6 30 6 yes 0.1283 4 1.8
7 50 6 yes 0.096 3 1.7
8 50 no enzyme yes 0.0941 2 1.5
9 50 6 no 0 2 N/A
F