Question

Define isoelectric point (pI)? How does it affect net charge on an amino acid? Calculate (show the calculations) the pI for histidine, and aspartic acid. pK values for these amino acids can be obtained from lab manual.

Answer 1

The isoelectric point (pI ), is the pH at which a particular molecule carries no net electrical charge.

If the pH of the buffer is above the pI of the protein being run, the protein will migrate to the positive pole (negative charge is attracted to a positive pole). If the pH of the buffer is below the pI of the protein being run, the protein will migrate to the negative pole of the gel (positive charge is attracted to the negative pole). If the protein is run with a buffer pH that is equal to the pI, it will not migrate at all. This is also true for individual amino acids.

Histidine pKa1 = 1.82, pKa2= 9.17, pKa3 = 6.00,

Isoelectric point = (pKa1 + pKa2} / 2

Isoelectric point = 7.59

Aspartic acid  pKa1 = 1.88, pKa2= 9.60, pKa3 = 3.65,

Isoelectric point = 2.77