TI  - Correlation of two-hybrid affinity data with in vitro measurements.
PG  - 5820-9
AB  - Since their introduction, the interaction trap and other two-hybrid
      systems have been used to study protein-protein interactions. Despite
      their general use, little is known about the extent to which the degree of
      protein interaction determined by two-hybrid approaches parallels the
      degree of interaction determined by biochemical techniques. In this study,
      we used a set of lexAop-LEU2 and lexAop-lacZ reporters to calibrate the
      interaction trap. For the calibration, we used two sets of proteins, the
        <prot>Myc</prot>  - <prot>Max</prot> - <prot>Mxi1</prot>  helix-loop-helix proteins, and wild-type and
      dimerization-defective versions of the lambda cI repressor. Our results
      indicate that the strength of interaction as predicted by the two-hybrid
      approach generally correlates with that determined in vitro, permitting
      discrimination of high-, intermediate-, and low-affinity interactions, but
      there was no single reporter for which the amount of gene expression
      linearly reflected affinity measured in vitro. However, some reporters
      showed thresholds and only responded to stronger interactions. Finally,
      some interactions were subject to directionality, and their apparent
      strength depended on the reporter used. Taken together, our results
      provide a cautionary framework for interpreting affinities from two-hybrid
      experiments.
AD  - Fox Chase Cancer Center, Philadelphia, Pennsylvania 19111, USA.
