Passage I

Enzymes catalyze (speed up) the rate of a biological reaction without being consumed in the reaction. Pyrococcus furiosus, an organism that lives in ocean thermal vents where temperatures remain near 100℃, produces the enzyme prolidase. Prolidase catalyzes the breakdown of proteins to release the amino acid proline. An enzyme's activity is a measure of its effectiveness for a given reaction expressed as a percent of its maximum measured effectiveness.

Experiment 1

Many enzymes function only in the presence of a metal ion, called a cofactor. One mL of a standard solution containing prolidase and 0. 4 mM (millimolar, a unit of concentration) cobalt ion (Co²⁺) was mixed with 1 mL of a solution of Protein X (of which proline is a component) at a pH of 7.0 and 100℃. After 10 min, a substance that reacts with free proline to produce a bright red color was added to the solution. The solution was diluted to 100 mL, and the intensity of the red color was measured and used to calculate the activity. The procedure was repeated for different concentrations of Co²⁺, and then for manganese ion (Mn²⁺) and zinc ion(Zn²⁺) at all of the concentrations (see Table 1).

Experiment 2

Experiment 1 was repeated, but Co²⁺ (held constant at 1.2 mM) was the cofactor in all trials, and the temperature was varied from trial to trial (see Table 2)

Experiment 3

Experiment 2 was repeated, but from trial to trial, temperature was held constant at 100℃, and pH was varied (see Table 3).