1What an enzyme does, and what it does not do

Glucose left in a beaker with oxygen will not burn at room temperature in your lifetime; inside a cell, a chain of enzymes releases its energy in seconds. Three things are true of every enzyme in this course:

1. It is a protein. Its function comes from its folded three-dimensional shape.
2. It speeds up a reaction; it does not cause one. An enzyme cannot make a reaction happen that would not happen without it. It only makes the reaction reach the same end point sooner.
3. It is not used up. After the products leave, the enzyme binds a new substrate molecule. So a cell needs only a little of each enzyme, and it can regulate a pathway by switching enzymes on and off (Topic 3.2) or making more or fewer of them (Unit 6).

2Activation energy

Before reactants can turn into products, their bonds have to be strained and partly broken. That takes an input of energy, even when the reaction as a whole releases energy, as a boulder must be pushed over a small hump before it rolls downhill. The height of the hump is the activation energy.

Figure 1 plots the energy in the molecules against the progress of the reaction. Read it in this order:

• The substrate starts higher than the products. The drop between them is the energy released, the same with or without the enzyme.
• The grey dashed curve, without the enzyme, has a tall hump. Its activation energy is the arrow from the substrate level to the top of that hump.
• The teal curve, with the enzyme, has a lower hump, so its activation energy is smaller.

With a lower barrier, a far larger share of substrate molecules have enough energy to react at any moment, so the reaction goes faster.

How? The active site holds the substrate in the right position, strains the bonds that must break, and surrounds it with charged or polar R groups that make the change easier. You need the principle, not the detail for any one enzyme.

An enzyme lowers the activation energy of a reaction. It does not change the energy of the reactants or the products, and it is not used up.

3The active site: shape and charge

The active site is a pocket on the enzyme's surface, lined by the R groups of a few amino acids that folding brings together. Two things decide whether a molecule can bind:

• Shape. The substrate must fit the pocket, as a key fits a lock.
• Charge. The R groups lining the site carry charges, partial charges or none. A substrate binds where its own charges are attracted to opposite charges in the site, and where its non-polar parts sit against non-polar R groups. Like charges repel.

Both have to match. When the substrate binds, the two form the enzyme–substrate complex. The active site is not rigid: binding makes it shift slightly to grip the substrate more closely (often called induced fit), which helps strain its bonds. The products form and are released, and the site returns to its original shape.

This is the enzyme–substrate complex model, and it explains specificity: each enzyme catalyses one reaction, because only its own substrate matches its active site.

Worked example: predicting the effect of a change. An invented enzyme has an active site lined by three amino acids. One of them has a positively charged R group, and the substrate carries a negatively charged phosphate group that sits against it. A mutation replaces that amino acid with one whose R group is non-polar.

• Claim: the enzyme will bind the substrate less well, so the rate of reaction will fall.
• Evidence: the positive R group attracted the negative phosphate; the new R group carries no charge.
• Reasoning: binding depends on a match of charge as well as shape. Without the attraction, fewer substrate molecules stay bound long enough to form the enzyme–substrate complex.

Claim, evidence, reasoning: AP free-response points are built on that pattern.

4Measuring how fast an enzyme works

The rate of an enzyme-catalysed reaction is how much product appears, or how much substrate disappears, per unit time. The formula sheet gives it as

An invented experiment. Catalase, an enzyme from liver and yeast, breaks hydrogen peroxide into water and oxygen. A student mixes catalase with hydrogen peroxide and collects the oxygen.

The rate over the first 20 seconds:

The rate over the last 20 seconds:

The rate falls six-fold because the substrate is being used up (not the enzyme): fewer hydrogen peroxide molecules collide with active sites each second. This is why experiments compare initial rates, measured before the substrate concentration has fallen much. Always write the unit.

5Designing an enzyme experiment

Four words carry the marks in questions on experimental design.

• Independent variable: what the experimenter deliberately changes. For example, the concentration of hydrogen peroxide.
• Dependent variable: what is measured to see the effect. For example, the volume of oxygen produced per second.
• Controlled variables: everything else that could affect the rate, kept the same: temperature, pH, the amount of enzyme.
• Control: a trial set up to show what happens when the factor being tested is absent or at a known value, so the results can be compared with it.

Two controls appear again and again in enzyme questions:

• A negative control shows what happens without active enzyme: the enzyme boiled first, or water in its place. If oxygen still appears, something other than the enzyme is causing the reaction.
• A positive control uses a condition known to work, to show the method can detect the reaction.

To justify a control, say what it rules out. "The boiled-enzyme tube is a control" earns nothing. "The boiled-enzyme tube shows that the oxygen in the other tubes was produced by active catalase and not by the hydrogen peroxide breaking down on its own" earns the point.

Repeat each trial at least three times so you can calculate a mean and a standard error.