The protein has a MW of 35,000. One is interested in determining the affinity of P for leonine. An amino acid binding protein (P) is released from E. coli by the osmotic shock, and purified to homogeneity. To do this, you choose to use equilibrium dialysis. A solution of protein (0.5mg/ml) is placed on one side of a compartment separated by a dialysis membrane. On the other side is buffer of equal volume with 4 x10-5 ML-Leucine14C, which is radio labeled with 14C. You find that the compartment containing protein had 2.3 X10-5 ML-Leucine14C (bound + free) after a lengthy equilibration. The compartment without had 1.7 X10-5 L-Lucien 14C (free).
1. Determine the concentration of bound L-Leucine14C.
2. Determine the concentration of free protein.
3. Determine the Kd, assuming one site per molecule.