🧬AP Biology
1. Enzymes are biological catalysts that speed up chemical reactions by lowering activation energy. The structure of an enzyme, particularly its active site, is critical for its function and is maintained by various chemical interactions.
Polyphenol oxidase (PPO) is an enzyme found in many plants that catalyzes the oxidation of phenolic compounds, leading to tissue browning. Researchers isolated PPO from a wild-type (WT) potato strain and a mutant strain (Mut-A) that exhibits rapid browning. To compare the stability of the two enzymes, the researchers measured the initial rate of the reaction catalyzed by each enzyme at various temperatures ranging from 10°C to 70°C.
The reaction mixtures contained saturating amounts of substrate and equal concentrations of enzyme. The relative activity was calculated by comparing the rate at each temperature to the maximum rate observed for the wild-type enzyme (Figure 1).
The researchers hypothesized that the mutation in Mut-A disrupts an ionic bond that stabilizes the enzyme's active site. To test this, they treated purified WT and Mut-A enzymes with increasing concentrations of sodium chloride (NaCl). NaCl dissociates into ions that can disrupt ionic bonds within proteins. The researchers measured the percent of initial activity remaining for each enzyme after incubation in 0 M, 0.5 M, and 1.0 M NaCl solutions (Figure 2).
Describe how the R-groups of amino acids contribute to the tertiary structure of a protein.
Figure 1. Relative activity (percent of the maximum wild-type rate) of polyphenol oxidase (PPO) from Wild-Type (WT) potatoes and mutant (Mut-A) potatoes measured at discrete temperatures from 10°C to 70°C. Reaction mixtures contain saturating substrate and equal enzyme concentrations. Error bars show ±SE.
Identify the dependent variable in the experiment shown in Figure 1.
Justify the researchers' decision to include the measurement of enzyme activity at 30°C for both the wild-type and mutant enzymes.
Based on Figure 1, describe the effect of the mutation on the optimal temperature of the enzyme.
Figure 2. Percent of initial PPO activity remaining after incubation in increasing NaCl concentrations for Wild-Type (WT) and Mut-A enzymes. Error bars show ±SE.
Identify the independent variable in the researchers' second experiment (data shown in Figure 2).
Based on Figure 2, identify the enzyme that demonstrates lower stability in the presence of NaCl.
The gene encoding the wild-type enzyme contains a coding region of 1,200 nucleotides. The gene for the mutant enzyme contains a deletion of 15 nucleotides in the coding region. Assuming the deletion does not cause a frameshift, calculate the difference in the number of amino acids between the wild-type and mutant proteins.
Researchers claim that the mutation in the mutant enzyme disrupts an ionic bond that stabilizes the protein structure. Using data from Figure 2, support the researchers' claim.
Justify the researchers' claim based on your understanding of how high concentrations of salt (NaCl) affect the chemical bonds within a protein's tertiary structure.
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