Enzymes: Catalysis and Specificity

• Enzymes are proteins that act as catalysts to speed up biochemical reactions in cells.
• Enzyme structure determines its specificity, allowing it to bind to specific substrates.
• Disruption of a protein’s shape, or denaturing, can prevent it from functioning.
• Enzymes are essential for cell function, aiding in metabolic processes like catabolism and anabolism.
• Homeostasis, the maintenance of constant internal conditions, is critical for optimal enzyme function.

Key Vocabulary

• Enzyme: A biological catalyst.
• Catalyst: A substance that speeds up a reaction without being consumed.
• Specificity: The ability of an enzyme to bind to a specific substrate.
• Homeostasis: Maintaining a stable internal environment.
• Catabolism: Breaking down complex molecules.
• Anabolism: Building complex molecules from simpler ones.
• Denaturing: Altering the shape of a protein, which can prevent it from functioning.
• Positive control: A test that demonstrates what a positive result should look like.
• Negative control: A test that demonstrates what a negative result should look like.
• Pepsin: A protease enzyme found in the stomach.
• Turbidity: The cloudiness of a solution.
• Calibration: Adjusting an instrument to ensure accurate measurements.

Introduction to Biomolecules and Enzymes

• Biomolecules, like carbohydrates, must be broken down (catabolized) into smaller subunits to be absorbed.
• Isomerase is an enzyme that breaks down fructose.
• Enzymes work only on specific substrates due to their unique shape and structure.
• Enzymes have an active site where the substrate binds.
• Lactose, but not fructose, is broken down when lactase is present.

Enzyme Function and Conditions

• Enzymes function best within a narrow range of conditions.
• Amylase functions best at neutral pH (6.7-7), at 37°C (98.6°F).
• Chemical reactions using enzymes convert complex molecules to smaller molecules.
• Anabolism, building complex molecules, relies on products from digestion.

Activity 1: Protease (pepsin) Enzyme Function

• The activity aims to test how changing conditions affects enzyme activity.
• Protease, commonly known as pepsin, catalyzes the catabolism of protein.
• Egg white slurry is used.
• Cloudiness reduction indicates protein breakdown.
• A turbidity meter measures solution cloudiness.
• Protein digestion occurs mainly in the stomach which has an acidic pH due to hydrochloric acid (HCl).
• Human stomach temperature stays constant around 37°C (98.6°F).
• A hypothesis should be constructed to explain what is needed to digest protein.
• Determine a prediction based on the hypothesis, by exposing egg white to pepsin under typical stomach conditions or altered conditions.
• Only change the enzyme, or concentration while testing the hypothesis.
• The independent variable is the factor being changed, while the dependent variable is the measured response

Experimental Controls

• Positive controls yield expected results; pepsin at low pH and body temperature mimics stomach conditions.
• Negative controls yield no results; pure water calibrates the turbidity meter to ensure accurate measurements

Safety and Procedure

• Wear safety glasses and gloves when working with chemicals and cleaning test tubes.
• Spilled HCl should be neutralized with baking soda.
• Each lab group should plan to adjust one independent variable.
• All test tubes must include egg albumin.

Experimental Setup

• A control reaction is run using known optimal conditions.
• One factor (enzyme, pH, or temperature) is changed in the experimental cuvette, while keeping all other factors like the control.
• The turbidity meter must be calibrated before use.
• Turbidity of control and experimental cuvettes is measured and recorded in NTU (Nephelometric Turbidity Units).
• Pre- and post-incubation turbidity measures are to be recorded.
• Observe and record any changes to the egg white and solution.

Analysis

• The biological rationale should link biomolecule and enzyme knowledge to the pepsin digestion experiment.
• A hypothesis and prediction should be created with properly graphed variables (independent vs. dependent).
• After collecting results, observations are discussed, and the hypothesis is either supported, refuted, or partially supported.
• Choose an appropriate graph with accurately labeled units.