Enzymes Central to Biochemical Processes: Aldolase Essay

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Enzymes are central to every biochemical process. Due to their high specificity they are capable of catalyzing hundreds of reactions that signifies their vast practical importance.

This experiment was conducted to investigate the enzymatic activity of the enzyme aldolase by using different urea concentrations. Stronger solutions of denaturant in this case urea were used to increase the polypeptide subunit unfolding and the dissociation of the tetramer.

Aldolase is an enzyme that consists of four polypeptide chains that are identical. The enzyme itself is commonly found in a rabbit muscle tissue where it is recruited to catalyse the breakdown of the fructose1,6-biphosphate to two products; dihydroxyacetone and glyceraldehyde
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The products produced by the first reaction were used as a substrate for the second. In this case the enzyme used NADH, which resulted in the decreased absorbance due to the NADH oxidation to NAD+. In addition, the spectrophotometer was used as a measuring device to follow the change in absorbance of the NADH molecules at 340nm.

Incorporation of assay controls included setting up a spectrophotomer and running the chart recorder with a full-scale deflection before the start of the assay. The set recorder had a corresponding value of 1 for the change in the absorbance. Therefore, prior testing was done to observe whether a change occurred in the readings. This helped to indicate that the results were valid, as they could have been affected by a fault during the setting up of the spectrophotometer. On the other hand this was considered as one of the controls for the experiment. Nevertheless, a new cuvette had to be used for each assay.
Also, the enzyme was treated with different urea concentrations before it was added to the assay mixture. The amount of the second and third enzymes was essential. Likewise, this would have had an effect on the rate of the measured aldolase activity. Hence, the concentrations for both enzymes was measured and calculated prior to the experiment whilst preparing the assay mixture. Furthermore, in order to measure thiol group reactivity the base line was adjusted to zero for each cuvette. This was done to make sure that the results were
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