Why did freezing have no effect on amylase activity in the described setup?

• A. Freezing has no effect on enzyme activity
• B. Freezing denatures the enzyme
• C. Freezing increases substrate concentration
• D. Freezing removes the substrate

Answer: (B)

Free enzyme activity depends on both temperature and the enzyme’s folded structure. Freezing can disrupt that structure because ice formation and changes in the solvent environment destabilize the noncovalent interactions (hydrogen bonds, ionic interactions, hydrophobic packing) that hold the enzyme in its active shape. When the active site loses its precise geometry, the substrate can no longer bind properly and catalysis stops, so amylase activity falls to essentially zero. In this setup, freezing would denature the enzyme and eliminate activity, which is why the choice describing denaturation best explains the result. Substrate concentration or presence isn’t increased or removed by freezing, so those ideas don’t account for the loss of activity.