MBB 400a/700a
Problem Set #2

1. You conduct an experiment to look at the interactions between protein X and ligand Y.  The beaker in which you conduct your experiment contains a semipermeable membrane that retains the protein to one side, but allows the ligand to pass through.  The concentration of protein X in half the beaker remains constant, [x] = 1.4e-5 M.  You add ligand Y to the beaker to produce the total Y concentrations as listed below and wait for the beaker to equilibrate.  You then measure the concentration of free Y by taking an aliquot out from the side without protein.  The data is listed below.  Figure
[Y] total (M)   [Y] free (M)
5e10-6           1.4e-6
1e-5               3.3e-6
1.68e-5          6e-6
2.44e-5          1e-5
5.16e-5          3e-5

What is the Kd of this interaction?  The Ka?  What is the maximum number of ligands bound per molecule of protein X?

2. The l repressor may be cleaved by trypsin into two domains, a dimerization domain and a DNA binding domain as shown in the following figure. Figure
You are studying the interactions of the l repressor with DNA. Sketch the binding curve for wild-type l repressor binding to DNA.
The following species were produced by trypsin cleavage of the repressor.  Sketch the binding curves for each of these cases.

3. For the following problem, you may consult the paper handed out in class.
Cunningham, B. C., Mulkerrin, M. G., and Wells, J. A., 1991.  Dimerization of Human Growth Hormone by Zinc.  Science 253, 545-548.
Estimate the percentage of hGH that exists in the human pituitary as a dimer.  If 10% of the pituitary gland contents were released into the bloodstream, what would be the percentage of hGH in dimer form?  Please state all of your assumptions.