🧬AP Biology
🧬AP Biology
FRQ 1 – Interpreting and Evaluating Experimental Results (Long)
Unit 1: Chemistry of Life
Practice FRQ 1 of 201/20
1. Proteins are essential macromolecules that perform a diverse range of functions in biological systems, including catalysis, transport, and structural support. The function of a protein is intimately tied to its specific three-dimensional shape.
Researchers isolated a digestive enzyme, Protease A, from a bacterium found in a hot spring environment. They also identified a mutant strain of the bacterium that produces a variant of the enzyme with a single amino acid substitution. To investigate the functional differences between the Wild Type and Mutant enzymes, the researchers purified both proteins and measured their relative catalytic activity across a range of temperatures from 20°C to 80°C. The assays were performed at a constant pH of 7.
In the activity assay, the enzyme was mixed with a specific substrate, and the rate of product formation was measured. A negative control containing substrate but no enzyme was also tested at each temperature and showed no product formation (data not shown). The results of the experiment are presented in Figure 1.
To investigate the structural stability of the enzymes, the researchers incubated purified samples of both the Wild Type and Mutant proteins with a generic protease (an enzyme that degrades other proteins) for 60 minutes at 40°C. Following incubation, the samples were analyzed using SDS-PAGE (gel electrophoresis) to separate the proteins by size. The intact Protease A protein has a molecular mass of 34 kDa. The results are visualized in Figure 2.
A. Describe how the R-groups of amino acids contribute to the tertiary structure of a protein.
Figure 1. Effect of temperature on the relative catalytic activity of Protease A (Wild Type vs Mutant) measured at pH 7. Error bars indicate ±SE (±5 percentage points at every temperature).
B.
i. Identify the dependent variable in the experiment shown in Figure 1.
ii. Justify the researchers' decision to include a negative control reaction containing no enzyme in the experiment described in Figure 1.
iii. Based on Figure 1, describe the difference in the optimal temperature between the Wild Type and Mutant enzymes.
Figure 2. SDS-PAGE diagram showing stability of Wild Type and Mutant Protease A after exposure to a generic protease for 60 minutes. Lanes include molecular mass markers (kDa) and enzyme samples at 0 min and 60 min.
C.
i. Identify the independent variable in the experiment shown in Figure 2.
ii. Based on Figure 2, identify the enzyme variant that demonstrates greater structural stability when exposed to the generic protease.
iii. The intact Protease A protein has a mass of 34 kDa. Based on Figure 2, the largest degradation fragment of the Wild Type enzyme in Lane 3 aligns with the 10 kDa marker. Calculate the difference in mass between the intact protein and this degradation fragment.
D.
i. Researchers claim that the Mutant enzyme is more resistant to unfolding (denaturation) than the Wild Type enzyme. Support this claim using evidence from Figure 2.
ii. The researchers determined that the mutation involves the substitution of a serine amino acid with a cysteine amino acid. Justify the claim that this substitution increases the stability of the protein's tertiary structure.
