Purification of Myoglobin (Mb). Purification tables are often used to keep track of the yield and purification of a protein. The specific activity is a ratio of the amount of the protein of interest, in this case Mb, obtained at a given step (mol or enzyme units) divided by the amount (mg) of total protein. The yield is the ratio of the amount of the protein of interest obtained at a given step (mol or enzyme units) divided by the original amount present in the crude extract, often converted to percent yield by multiplying by 100. The fold purification is the ratio of the specific activity of the purified protein to that of the crude preparation.
(a) For the purification table below, calculate the specific activity, % yield, and fold purification for the empty cells.
(b) Which step-DEAE or affinity chromatography-causes the greatest loss of Mb?
(c) Which step causes the greater purification of Mb?
(d) If you wanted to use only one purification step, which technique would you choose?