Pre-Lab Questions Lab 2 PDF

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Pre-lab questions for the second in person lab, Enzyme Kinetics. ...

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Enzyme Kinetics Pre Lab 2 Questions 1. How do enzymes speed up reactions? Enzymes speed up reactions by lowering the activation energy. When the substrate is positioned in a specific manner, the bonds they possess become unstable and thus reactive. The activation energy brings the substrate molecule into a state of transition. The enzyme is responsible for stabilizing the transition state. This in turn, causes the reaction to speed up as less energy is needed for the substrate to be converted into a product. 2. What enzyme are we using for this week’s lab and what is its substrate? How is this related to biofuel production? The enzyme we are using in this week’s lab is cellobiase. Cellobiase is a molecule composed of long chains of glucose. Cellobiase is involved in the last step of breaking down cellulose into glucose. Cellulose is the natural substrate of cellobiose. The cellulose is broken down from plants into sugar. This is vital in creating ethanol as well as other biofuels. The substrate used in the lab is p-Nitrophenyl which reacts to form p-Nitrophenol.

3. All enzymes have a maximal rate they can catalyze a reaction at. How can you determine the maximum rate? How would you collect that data? When adding enzyme to the artificial substrate, p-Nitrophenyl glucopyranoside, the maximum rate can be calculated by measuring the amount of p-Nitrophenol produced over time. An indicator is used to turn the color when the reaction takes place. The “yellowness” of the solution is directly related to the amount of p-Nitrophenol present. Additionally, a spectrophotometer can be used to give a quantitative value of the amount of yellow color present. The absorbance of values can be plotted to show the standard curve, thus the concentration. V0 =

V max × [S] (K m + [S ])

4. If a cell wants to increase a specific biochemical reaction, it can most easily change the concentration of the enzyme inside the cell. How does this affect the rate of reaction? Increasing the concentration of an enzyme inside the cell greatly increases the maximum reaction rate. An enzyme works at the maximum rate when the active site is completely saturated with the substrate. The initial rate of reaction can be determined experimentally when there are different starting concentrations used.

5. On the graphs provided, draw your predictions for how cellobiase enzyme would work under increasing temperature, pH, substrate concentrations, and enzyme concentrations.

Plot the reaction rate (product produced/time) on the y-axis vs. the increasing variable (low to high)....