The effect of catalase concentration on the rate of decomposition of hydrogen peroxide

Enzymes are biological catalysts that catalyze biochemical reactions in living cells and remain chemically unchanged at the end of the reaction. In an enzyme-catalyzed reaction, the substrate binds to the active site and forms enzyme-substrate complex with the enzyme. The enzyme then breaks the bonds in the substrate. The product of the reaction then leaves the enzyme, which remains unchanged after the reaction.

Figure 1. Free substrate colliding with a free enzyme, resulting in the catabolic reaction.

Enzymes work by lowering the activation energy for a reaction, thus increasing the rate of the reaction. Thus, as the rate or enzyme concentration increases, the rate of reaction increases, assuming that enzyme concentration is a limiting factor. Due to the three-dimensional shape of the enzyme, its active site is specific and only acts on a particular substrate.

In the liver, Hydrogen peroxide is a toxic by-product of fatty acid oxidation. When it is left alone, it is relatively stable and thus, each molecule of hydrogen peroxide can stay in the body for a few years. As hydrogen peroxide is active and harmful to cells and tissues of organisms, its decomposition therefore needs to be speeded up greatly in order to prevent it from intoxication in the cell.

Catalase is a common enzyme found in nearly all living organisms exposed to oxygen that catalyzes the decomposition of hydrogen peroxide to water and oxygen. It is one of the fastest acting enzymes it needs to maintain low levels of hydrogen peroxide. It is a very important enzyme in reproductive reactions. Likewise, catalase has one of the highest turnover numbers of all enzymes; one catalase molecule can convert millions of molecules of hydrogen peroxide to water and oxygen each second.

Reaction catalysed by catalase:

2 H2O2 ? 2 H2O + O2

While the complete mechanism of catalase is not currently known, the reaction is believed to occur in two stages:

H2O2 + Fe(III)-E ? H2O + O=Fe(IV)-E(.+)

H2O2 + O=Fe(IV)-E(.+) ? H2O + Fe(III)-E + O2

Here Fe()-E represents the iron center of the heme group attached to the enzyme. Fe(IV)-E(.+) is a mesomeric form of Fe(V)-E, meaning that iron is not completely oxidized to +V but receives some “supporting electron” from the heme ligand. This heme has to be drawn then as radical cation (.+).

As hydrogen peroxide enters the active site, it interacts with the amino acids Asn147 (asparagine at position 147) and His74, causing a proton (hydrogen ion) to transfer between the oxygen atoms. The free oxygen atom coordinates, freeing the newly-formed water molecule and Fe(IV)=O. Fe(IV)=O reacts with a second hydrogen peroxide molecule to reform Fe(III)-E and produce water and oxygen.

Catalase is a tetramer of four polypeptide chains, each over 500 amino acids long. It contains four porphyrin heme (iron) groups that allow the enzyme to react with the hydrogen peroxide. The optimum pH for human catalase is approximately 7, and has a fairly broad maximum (the rate of reaction does not change appreciably at pHs between 6.8 and 7.5). The pH optimum for other catalases varies between 4 and 11 depending on the species. The optimum temperature also varies by species. Catalase is usually located in a cellular, bipolar environment organelle called the peroxisome. Peroxisomes, which get their name from peroxide and hydrogen, are spherical, 0.3 – 1.5 mm in diameter and bounded by a single membrane.

The factors affecting rate of decomposition are temperature, pH and changes, concentration of hydrogen peroxide and concentration of catalase. As this investigation investigates the effect of concentration of catalase on the rate of decomposition of hydrogen peroxide, the temperature, pH and concentration of hydrogen peroxide are kept constant throughout the experiment.

Hypothesis

Based on the background information, I hypothesize that the general trend will be such that as the concentration of catalase increases due to the number of potato discs increasing, the rate of decomposition of hydrogen peroxide will also increase and this is translated into the mean air pressure measured to be increasing. However, the hypothesized graph will have a concave down as seen in figure 2. This is because as the concentration of catalase increases, more enzyme-substrate complexes will be formed and there will be fewer hydrogen peroxide molecules that are free, thus causing the rate of reaction to increase at a slower rate.

It will reach a point whereby all the hydrogen peroxide molecules are being decomposed such that even as the concentration of catalase increases, the rate of reaction will not increase. Concentration of catalase will no longer become a limiting factor but instead, concentration of hydrogen peroxide solution becomes a limiting factor. Rate of reaction can now only increase if concentration of hydrogen peroxide solution increases.