Enzyme Kinetics of Beta-Galactosidase

Topics: Enzyme, Catalysis, Chemistry Pages: 5 (1705 words) Published: July 10, 2011
This experiment is to study and measure the enzyme activity of β-galactosidase in the different concentrations of o-Nitrophenylgalactoside (ONPG) using a spectrophotometer. The spectrophotometer was also set at 420nm, a wavelength which is best for recording the absorbance values for the experiment. From the results, 0.9mM ONPG solution has the highest absorbance and 0.1mM ONPG solution has the least. Also, 0.5mM ONPG solution has the highest rate of enzyme activity and it is the most efficient as the enzyme activity of the ONPG solution continues even though the other concentrations of ONPG solution has already stopped the enzymatic reactions as the substrate is already used up.

This experiment is to study and measure the enzyme activity using a spectrophotometer and to understand the practical aspects of handling enzymse. This is done when the colourless ONPG is split into galactose and o-Nitrophenol (a yellow compound). The higher the enzyme concentration, the higher the absorbance value. Different molecules absorb different wavelengths of light. For this experiment, the spectrophotometer is set at 420nm, so that we can obtain the best absorbance results. ONPG →galactose+ o-Nitrophenol

A catalyst is a substance that reduces the activation energy of a chemical reaction, making it energetically viable. It is also used to speed up the rate of a chemical reaction. Enzymes are an example of a catalyst that can be found in the body. They are biological catalysts which are mainly made up of proteins. It is produced to speed up chemical reactions and remain unchanged after a reaction. Enzymes have active site for the substrate to attach to, either to be broken up or joined together. They are also specific in their reactions, they only speed up certain reaction as the active site can only fit a certain substrate and does not work for the other substrates. Also, enzymes will only work properly upon strict optimum conditions. They lower the activation energy for a reaction, thus requiring lesser energy for the reaction to occur, therefore increasing the speed of reaction.

A substrate is a molecule upon which an enzyme acts on. Enzymes are used to catalyse chemical reactions and the substrate will bind to the active site of the enzyme and an enzyme-substrate complex is formed. The substrate is then transformed into one or more products. Also, two or more substrates can bind to the active site of the enzyme and the substrates are then transformed into a different product. There are many factors that affect the rate of enzyme activity – temperature, pH, enzyme concentration, substrate concentration and the presence of any inhibitors or activators. The changes in pH can lead to the breaking of ionic bonds that hold the tertiary structure of the enzyme, thus it loses its active site, and therefore the substrate can no longer fit into it. Thus, there will be a decrease in enzyme activity. Each enzyme has its own optimum pH and it can vary considerably depending on the environment the enzyme works best in. The increase in temperature increases the rate of enzyme activity as there are more collisions between the substrate and the enzyme, thus forming more enzyme-substrate complex. However, the enzyme activity will start to decrease when the temperature rises above the optimum temperature for the enzyme. This is because denaturation of the enzyme is taking place and the active site is lost, therefore leading to lesser enzyme-substrate complex formed.

The concentration of the substrate can affect the speed of enzymatic reactions. As substrate concentration increases, the speed of the reaction increases until a point where all enzymes are used for reactions and the increment of the substrate concentration will have no effect on the rate of reaction. The increase of the enzyme concentration can increase the rate of enzyme activity if there are more substrate molecules than enzyme molecules. As there are...
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