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Heat Transfer Write Up For Engineering Cwk
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Date : 18/11/2012
Author Information
Uploaded by : Tanvir
Uploaded on : 18/11/2012
Subject : Engineering
Introduction ?-galactosidase is a large class of enzymes usually found in the bacteria Escherichia coli, which is in the gut. It has an optimum pH of around 6-7 and an optimum temperature of around 37°C, although these can vary.
As with all enzymes ?-galactosidase is a protein with a tertiary structure. Different types of ?-galactosidase enzymes have different structures mainly shown in the active site. This enables them to be specific to their substrate for example; the ?-galactosidase enzyme lactase can only catalyse the hydrolysis of lactose.
?-galactosidase catalyses the hydrolysis of ?-D-galactosides into monosaccharides. For example, it can catalyse the hydrolysis of lactose into glucose and galactose. It can also catalyse the hydrolysis of o-nitrophenol-?-D-galactoside (ONPG) into o-nitrophenol (ONP) and galactose.
?-galactosidase can be obtained from many different sources such as animals, plants, microorganisms etc. The most common way of obtaining ?-galactosidase is from microorganisms. This is because microorganisms offer various advantages over other sources. Some of these are easy handling, higher multiplication rate and higher production yield. However, the purification cost is a major disadvantage and a lot of research is going into reducing it. Also, due to the moderate stability of the enzyme being a limitation, there is a need to use different methods for stabilization. This is done by immobilization (Panesar, et al., 2010). Therefore, most ?-galactosidase produced industrially is immobilized.
?-galactosidase has many important applications. The main use is in the food industry. For example, in the dairy industry requires it to ferment sugars from lactose. The enzyme is also isolated from bacterial systems and added to dairy products to breakdown the remaining lactose. The product becomes lactose-free and hence does not cause digestive disruption to those suffering from lactose intolerance (Hensley, 2008).
In this experiment, ONPG was hydrolysed to ONP with the catalysis ?-galactosidase. The light absorbance was measured. A Lineweaver-Burk plot and Eadie-Hofstee plot were made at the end. This report shows all of this and also compares both of graphs.
Discussion Experimental data: Due to an experimental error, all the v/[s] values were 0.33 recurring so the Eadie-Hofstee plot was not drawn for this data. The thermostatic water bath did not remain at a constant 37°C but instead kept on increasing. However, Km and Vmax were both found from the Lineweaver-Burk plot. The Km and Vmax values calculated from the graph are accurate as the values found from the intercepts and gradient are close. For example, when calculated from the X-intercept, Km is 22.22mmol/mL and when calculated from the gradient, it is 22.32mmol/mL. These values are very close and show the graph plotted is accurate. The small difference could have been due to various experimental errors such as the faulty water bath. It could also have been due to the precision of the graph, the intercept is harder to read with less precision.
The reliability of the experiment could have been improved in various ways. The thermostatic water bath could be checked by testing it before actually using it. Also, the time intervals used to place the test tubes into the water bath could have been increased. This would allow more time to get ready for the next test tube to be placed in. In addition, the amount of repeats could have been increased. This would make the results much more reliable as the value for absorbance will be more accurate.
Data provided: The values of Km and Vmax for the Lineweaver-Burk plot vary depending on which method is used to find them. However, these values are still close to each other. The Km and Vmax values for the Eadie-Hofstee plot stay the same regardless of the method used to find them. This shows the Eadie-Hofstee plot gives a more reliable and accurate value for Km and Vmax.
The Km value obtained from both graphs was 333.33µmol/L with the exception of the value obtained with the gradient method on the Lineweaver-Burk plot. The different value from the gradient method was due to the Vmax value used on the Lineweaver-Burk plot. The Vmax values obtained from both graph are different but they are still very similar to one another. The difference could have been because in the Lineweaver-Burk plot the data is less evenly spread than in the Eadie-Hofstee plot and so the line of best fit is harder to determine for the Lineweaver-Burk plot. Also, reading the values off the graph was difficult so a more sensitive scale may need to be used.
Conclusion The values for Km and Vmax obtained from both graphs were similar. However, the Eadie-Hofstee plot had more consistent values showing it to be more reliable and accurate than the Lineweaver-Burk plot. However, the Eadie-Hofstee plot is very sensitive and a small experimental error could alter it greatly.
In conclusion, the differences in the values for Km and Vmax are mainly due to experimental and human errors. Both types of plots give very similar values for Km and Vmax. However, if the experiment is done perfectly the Eadie-Hofstee plot gives the more reliable values as are more consistent.
Bibliography Hensley, A.L., 2008. The Enzyme Beta-Galactosidase: Structure and Function, [online] Available at: http://chemistry.berea.edu/~biochemistry/2008/ah/ [Accessed 09 January 2012]
Panesar, S.P., Kumari S. and Panesar, R., 2010. Potential Applications of Immobilized ?-Galactosidase in Food Processing Industries, [online] Available at: http://www.hindawi.com/journals/er/2010/473137/ [Accessed 09 January 2012]
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