Revise Factors that affect enzyme action for Cambridge 9700 (3.2)

Temperature is a critical factor influencing the kinetic energy of molecules. As temperature increases from low levels, both enzyme and substrate molecules gain more kinetic energy. This leads to more frequent and energetic collisions, increasing the likelihood of forming enzyme-substrate complexes and thus, the rate of reaction. For every 10°C rise in temperature, the rate of reaction approximately doubles, a concept known as the Q10 temperature coefficient.

This trend continues up to the optimum temperature, the temperature at which the enzyme exhibits maximum catalytic activity (typically around 37-40°C for human enzymes). Beyond this optimum, the increasing kinetic energy causes vigorous vibrations within the enzyme molecule. These vibrations break the weak hydrogen and ionic bonds that maintain the specific tertiary structure of the protein. This causes a permanent change in the shape of the active site, a process called denaturation. A denatured enzyme can no longer bind to its substrate, causing a rapid and irreversible decrease in the reaction rate.

The pH of the environment, a measure of hydrogen ion (H⁺) concentration, significantly impacts enzyme activity. The R-groups of the amino acids that make up the enzyme can be acidic or basic. Changes in pH alter the concentration of H⁺ and OH⁻ ions, which can interact with these R-groups, changing their charges. This disrupts the ionic and hydrogen bonds that maintain the enzyme's specific tertiary structure, particularly the shape of the active site.

Each enzyme has an optimum pH at which its active site has the precise conformation for maximum catalytic efficiency. Deviations from this optimum pH cause the active site to change shape, reducing its affinity for the substrate and lowering the reaction rate. Extreme changes in pH (highly acidic or highly alkaline) can cause irreversible denaturation. For example, pepsin in the stomach works optimally at pH 2, while trypsin in the small intestine functions best at pH 8.

With a fixed enzyme concentration, increasing the substrate concentration initially increases the reaction rate. At low substrate concentrations, many active sites are unoccupied, and the rate of reaction is limited by how often a substrate molecule collides with an active site. As more substrate is added, collision frequency increases, more enzyme-substrate complexes form per unit time, and the reaction accelerates.

However, this increase is not indefinite. Eventually, a point is reached where all available active sites are occupied (saturated) with substrate molecules. The enzyme is working at its maximum capacity. At this saturation point, adding more substrate will not increase the reaction rate further. The rate plateaus at its maximum velocity (Vmax). The enzyme concentration now becomes the limiting factor.

Assuming there is an excess of substrate (i.e., substrate is not a limiting factor), increasing the enzyme concentration will directly and proportionally increase the rate of reaction. With more enzyme molecules present, there are more available active sites to bind with the substrate. This allows more enzyme-substrate complexes to be formed simultaneously, leading to a faster rate of product formation. If you were to plot reaction rate against enzyme concentration (with excess substrate), you would see a straight line passing through the origin. However, if the substrate concentration becomes limited, the rate will plateau as there isn't enough substrate to occupy all the available active sites.

Enzyme inhibitors are substances that bind to an enzyme and reduce its activity. This inhibition can be reversible or irreversible, and is a key mechanism for regulating metabolic pathways in cells.

a) Competitive Inhibition A competitive inhibitor has a molecular shape that is similar to the substrate. It competes with the substrate for the same active site. When the inhibitor is bound to the active site, the substrate cannot bind, and no reaction occurs. The level of inhibition depends on the relative concentrations of the inhibitor and the substrate. Increasing the substrate concentration can overcome competitive inhibition, as it increases the probability of the substrate binding to the active site instead of the inhibitor. Competitive inhibitors increase the apparent Km (more substrate is needed to reach ½ Vmax) but do not change the Vmax.

b) Non-competitive Inhibition A non-competitive inhibitor binds to the enzyme at a location other than the active site, known as an allosteric site. This binding causes a conformational change in the enzyme's overall structure, which alters the shape of the active site. As a result, the substrate can no longer bind effectively, or the enzyme is less efficient at catalysing the reaction. Since the inhibitor does not compete for the active site, increasing the substrate concentration cannot overcome this type of inhibition. Non-competitive inhibitors reduce the Vmax (as they effectively reduce the concentration of functional enzymes) but do not change the Km.