In the citric acid cycle, an enzyme oxidizes malate to oxaloacetate, with the production of NADH and the release of H+. You are studying this reaction using a suspension of bean cell mitochondria and a blue dye that loses its color as it takes up H+. You set up reaction mixtures with mitochondria, dye, and three different concentrations of malate (0.1 mg/L, 0.2 mg/L, and 0.3 mg/L).
Which of the following graphs represents the results you would expect, and why?

Verified step by step guidance

1. Understand the reaction: Malate is oxidized to oxaloacetate in the citric acid cycle, producing NADH and releasing H+. The blue dye loses its color as it takes up H+, meaning the reaction's progress can be tracked by the decrease in color intensity of the dye.
2. Interpret the experimental setup: The reaction mixtures contain mitochondria, dye, and varying concentrations of malate (0.1 mg/L, 0.2 mg/L, and 0.3 mg/L). Higher malate concentrations should lead to faster reaction rates, as more substrate is available for the enzyme to act upon.
3. Analyze the graphs: Graph (a) shows increasing color intensity over time, which does not align with the expected decrease in dye color as H+ is released. Graph (b) shows a decrease in color intensity, but all lines converge at the same endpoint, which does not reflect the effect of varying malate concentrations. Graph (c) shows a decrease in color intensity, with steeper slopes for higher malate concentrations, which matches the expected results.
4. Explain the expected results: As malate concentration increases, the reaction rate should increase, leading to faster uptake of H+ by the dye and a more rapid decrease in color intensity. This is consistent with the pattern shown in graph (c).
5. Conclude: Graph (c) represents the expected results because it shows a decrease in color intensity over time, with the rate of decrease proportional to the malate concentration.

Key Concepts

Citric Acid Cycle

The citric acid cycle, also known as the Krebs cycle, is a series of enzymatic reactions that occur in the mitochondria, where acetyl-CoA is oxidized to produce energy. This cycle generates electron carriers, such as NADH and FADH2, which are crucial for the electron transport chain. Understanding this cycle is essential for analyzing how malate is converted to oxaloacetate and the implications for energy production.

Enzyme Kinetics

Enzyme kinetics studies the rates of enzyme-catalyzed reactions and how they change in response to varying substrate concentrations. In this experiment, the concentration of malate affects the rate of the reaction catalyzed by the enzyme, which can be observed through the production of NADH and the consumption of H+. This concept is vital for predicting how changes in substrate concentration will influence the reaction's progress and the resulting graph.

Colorimetric Assay

A colorimetric assay is a method used to measure the concentration of a substance by determining the intensity of its color. In this experiment, the blue dye serves as an indicator of H+ concentration, changing color as it binds to protons released during the oxidation of malate. Understanding how the dye's color change correlates with the reaction's progress is crucial for interpreting the expected results and the corresponding graph.