LAB TWO Enzyme Catalysis

Background Concepts: Enzyme Catalyst Catalysis Catalase Substrate Product.

LAB TWO Enzyme Catalysis Background Concepts: Active site Reaction rate Initial reaction rate Free energy & enzyme activity Temp, pH, [E], [S] & initial rxn rate.

LAB TWO Enzyme Catalysis Background Concepts: Salt concentration pH Temperature Activators Inhibitors competitive noncompetitive.

LAB TWO Enzyme Catalysis Background Concepts: catalase 2H 2 O 2 2H 2 O + O 2

Materials: H 2 O 2, 1.5% Catalase solution Potato, macerated H 2 SO 4, 1.0M KMnO 4. LAB TWO Enzyme Catalysis

EXERCISE 2A: Test of Catalase Activity Take 10 ml H 2 O 2 Add 1 ml catalase solution Observe Repeat, boiling catalase first Repeat, using potato instead of catalase.

LAB TWO Enzyme Catalysis EXERCISE 2B: The Base Line Assay Take 10 ml H 2 O 2 Add 1 ml H 2 O (not catalase) Add 10 ml H 2 SO 4 and mix Titrate with KMnO 4.

5H 2 O 2 + 2KMnO 4 + 3H 2 SO 4 KSO 4 + 2MnSO 4 + 8H 2 O + 5O 2 EXERCISE 2B: The Base Line Assay LAB TWO Enzyme Catalysis

5H 2 O 2 + 2KMnO 4 + 3H 2 SO 4 KSO 4 + 2MnSO 4 + 8H 2 O + 5O 2 The amount of 2KMnO 4 used is proportional to amount of H 2 O 2 remaining in the sample. EXERCISE 2B: The Base Line Assay LAB TWO Enzyme Catalysis

EXERCISE 2C: The Uncatalyzed Rate of H 2 O 2 Decomposition Take 15 ml H 2 O 2 Store uncovered at room temperature for 24 hours Repeat part B.

LAB TWO Enzyme Catalysis EXERCISE 2D: The Enzyme Catalyzed Rate of H 2 O 2 Decomposition Reestablish the baseline Take 7 more 10 ml samples of H 2 O 2 We will allow the reaction to proceed for 10, 30, 60, 90, 120, 180, and 360 seconds.

LAB TWO Enzyme Catalysis EXERCISE 2D: The Enzyme Catalyzed Rate of H 2 O 2 Decomposition Take 10 ml H 2 O 2 Add 1 ml catalase extract Swirl gently for exactly X seconds Add 10 ml H 2 SO 4 and mix Titrate with KMnO 4 Repeat with other timings..