Project #15992 - An investigation into the activity of serum alkaline phosphatase

Laboratory write up.

EXERCISE 1: Determination of the optimum pH for alkaline phosphatase.

 

1.            Convert the highest absorbance reading obtained (which should correspond to the pH optimum) to micromoles product formed during the reaction, using the calibration graph of absorbance vs. concentration for 4-nitrophenol, provided.  (10 marks)

 

 

 

2.            Plot a graph on the provided sheet of absorbance (Y axis) vs. pH to show the optimum pH range for the enzyme. (10 marks)

 

EXERCISE 2: determination of total serum protein using the biuret assay

Results:

3.            Plot a graph on the provided sheet of absorbance (Y axis) vs. weight of protein present (mg)

(10 marks)

 

 

4.            Use this graph to read off the total protein present in 0.05 ml serum (sample 4). You will need this value to calculate the specific activity of the enzyme sample. (10 marks)

 

EXERCISE 3:

 

5.            Calculate the enzyme specific activity = A / P µmol.min-1.mg-1    (20 marks)

 

 

6.            Convert the specific activity value to SI units (mol.s-1.kg-1).      (10 marks)

 

 

 

 

 

7.            Why and how do pH changes affect enzyme structure and activity? (10 marks)

 

 

 

 

8.            In any determination of enzyme activity, the concentration of substrate must be such that the enzyme is working at a maximum efficiency i.e. the rate of reaction is enzyme limited and not substrate-limited. How could you demonstrate that this is the case in exercise 1? (20 marks)

 

 

Subject Science
Due By (Pacific Time) 11/05/2013 08:00 am
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