Two-hybrid system: yeast LexA-BD/B42-AD

Source was yeast cell line; bait produced as transgenic fusion protein; prey produced as transgenic fusion protein (prey from embryo cDNA expression library).

Two-hybrid system: yeast LexA-BD/B42-AD

Yeast cells also expressed constitutively active c-Src kinase domain, which was necessary for detection of the interaction.

Source was yeast cell line; bait produced as transgenic fusion protein; prey produced as transgenic fusion protein (prey was previously cloned reagent).

The requirement for activated Src kinase suggests that drpr must be phosphorylated to bind shark.

Antibody to drpr protein (not the HA tag) used for immunoprecipitation.

Source was cell extract of S2 cell line; bait produced from transfected construct; prey produced from transfected construct.

Source was cell extract of S2 cell line; bait produced from transfected construct; prey produced from transfected construct.

Both drpr isoforms tested, I and II, bind.

Shark and Src42A were inserted into rhodamine labeled liposomes and BG505 tagged Shark was added. A Shark-drpr interaction was inferred from quenching of Shark\'s donor fluorescence upon addition of ATP, presumably due to Shark binding to phosphorylated drpr bringing it into proximity with the rhodamine labeled liposome.

Interaction in vitro; fluorescence donor produced as a recombinant fusion protein in bacterial system; fluorescence acceptor produced as a recombinant fusion protein in bacterial system.