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SLE222 Biochemistry- Biochemical fractionation of milk proteins

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1a) Produce a figure using the image of the SDS-PAGE gel that you created in the prac. In your figure present only the lanes for the molecular mass standard, the skim milk sample and the whey sample. 1b) Provide a short description of the difference in the protein content between the skim milk and whey protein samples. Describe the proteins in terms of their size and provide a conclusion about the size of the casein proteins present in the milk sample. Must be less than 100 words 2a) Using data from the SDS-PAGE standards, create a standard curve. To do this, draw a graph of the log mole...

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Question Preview:

1a) Produce a figure using the image of the SDS-PAGE gel that you created in the prac. In your figure present only the lanes for the molecular mass standard, the skim milk sample and the whey sample. 1b) Provide a short description of the difference in the protein content between the skim milk and whey protein samples. Describe the proteins in terms of their size and provide a conclusion about the size of the casein proteins present in the milk sample. Must be less than 100 words 2a) Using data from the SDS-PAGE standards, create a standard curve. To do this, draw a graph of the log molecular mass of the different protein standards on the y axis against the migration distance of the proteins within the polyacrylamide gel on the x axis. See figure 13 on page 97 of your text for an example. Using Excel, create a scatter plot and generate a linear regression and straight line equation. Express the equation in terms of log molecular mass and distance migrated. Give the graph a title and write a figure legend, which would explain the graph. The figure legend should allow the reader to understand the graph. 2b) Using the straight line equation of the standard curve, calculate the migration distance of a 43 kDa protein. 3) You are studying a protein that you think consists of a heterodimer composed of a 30 kDa and a 40 kDa subunit, which are joined together by disulphide bonds. In addition to your protein sample you have available a SDS-PAGE gel, an electrophesis apparatus like the one you used in the laboratory, and a Bio-Rad Precision Protein Kaleidoscope standard (Bio-Rad catalogue number 161-0375). You also have access to two different sample buffers. Buffer A contains 4% (v/v) SDS and 5% (v/v) ß-mercapto ethanol. Buffer B contains 4% (v/v) SDS but no ß-mercapto ethanol. As per the practical, samples are heated at 95oC for 4 minues before they are loaded onto an SDS-PAGE gel and electrophoresed. 3a) Describe the experimental methods that would allow you to test your hypothesis. Provide sufficient detail such that an interested reader could faithfully repeat your experiment. In your answer describe the chemistry and biochemistry involved in your treatment. Must be less than 200 words 3b)Create a hypothetical SDS-PAGE gel showing predicted protein bands that you would observe in the experiment described above. Label these bands with the predicted sizes. This can be created within PowerPoint or any other software package. Add black bars (resembling protein bands) to the gel image below approximately where you would expect protein bands to migrate following electrophoretic separation of the sample. Indicate the sizes of the bands on the hypothetical gel. A figure legend is not required.

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Skim milk is composed of both casein protein and whey. Skim milk has higher casein protein content while compared to whey which has more whey protein. The proteins that are generally present in skimmed milk are “casein (25-35 kDa), alpha and beta lactoglobulin of 14 and 18 kDa  respectively.”(Livney Yoav D., 2010). Skim milk has approximately 34-45% of protein and casein is the major protein present.There is a size range for casein protein as it depends upon the temperature and pH of milk. If there is alteration in milk pH it will alter the internal structure of casein protein. 

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