Cooperativity in Repressor Binding and its Measurement

Cooperativity in Repressor Binding and its Measurement

As seen in the preceding section, repressor does not bind with equal affinity to the three operators on the right. More detailed experiments also reveal that repressor molecules do not bind independently to the

Table 14.4 The Binding Energies of Repressor Di-mers to the Various Operators and the Interaction Energies that Generate Cooperative Binding of Re-pressor Dimers

three operators (Table 14.4). Not surprisingly, repressors bound at the operators interact with one another and change the overall binding energy of repressor for an operator. Most unexpectedly, however, the results show that a repressor molecule bound at the middle operator can interact either with a repressor bound at O_R1 or with a repressor bound at O_R3, but it does not simultaneously interact with both. This is termed a pairwise interaction.

Operator mutants were used to measure the intrinsic affinity of repressor for the three operator sites as well as to reveal interactions between adjacently bound repressor molecules. By eliminating re-pressor binding to O_R2, the binding energy of repressor to O_R1 and to O_R3could be measured without complications caused by interactionswith adjacent repressors. Similarly, eliminating binding at O_R1 or O_R3 permitted determination of the other binding energies. Ultimately, this series of measurements provided enough data to permit calculation of the interaction energies of adjacent repressor molecules. The measure-ments showed that repressor binding at O_R2 has a dramatic effect on the binding of adjacent repressors.