The purpose of this unit was the kinetic analysis of Alcohol dehydrogenase under ideal conditions and in the presence of an inhibitor. In a normal reaction, the reactant is converted to the product, whereas in an enzyme-catalyzed reaction, the substrate reversibly binds with the enzyme to form the enzyme-substrate complex, which is then converted to the product and the free enzyme. The rate of the reaction is the concentration of substrate consumed or the concentration of product formed, therefore, the rate of an enzyme-catalyzed reaction is the velocity at which the product is formed from the substrate over time. There are many factors that influence enzymatic reactions, but two major ones are the enzyme and substrate concentrations. The ideal conditions for studying enzyme activity are at zero order reaction conditions as the rate of the reaction would be indicative of solely the characteristics of the enzyme. An important aspect of enzyme-catalyzed reactions is saturation, where at low substrate reactions, the rate of the reaction increases as the substrate concentration is increased. Eventually, there will reach a point where the enzyme is performing at the maximum activity so the reaction rate is no longer dependent on the substrate concentration …show more content…
Furthermore, NADH has an absorbance max at 340 nm, which allows for the reaction to be followed by the increase in A340 as NAD+ is reduced. From this experimental data, the initial velocity of the reaction can be obtained as this is the rate of the reduction reaction, the slope of the absorbance vs. time curve. Subsequently, this can be converted to the concentration per time using the Beer-Lambert equation. From the values obtained, the Michaelis-Menten and Lineweaver-Burk plots can be created, which would allow for the determination of the KM, Vmax, and