Interaction in vitro; bait produced as a recombinant fusion protein in bacterial system; prey produced and labeled by in vitro translation.

Both halves of gro are required for full interaction with en.

The en repressor domain was fused to a portion of h (aa 1-286) that is unable to bind gro.

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

Four of 38 independent clones identified using en as two hybrid bait corresponded to gro.

Two-hybrid system: yeast GAL4-BD/GAL4-AD

Interaction in vitro; bait produced as a recombinant fusion protein in bacterial system; prey produced and labeled by in vitro translation.

Positive control.

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

Two-hybrid system: yeast GAL4-BD/VP16-AD

Positive control.

Interaction in vitro; bait produced as a recombinant fusion protein in bacterial system; prey produced and labeled by in vitro translation.

Mutations in gro that disrupt binding to the eh1 motif (FxIxxIL) typically also disrupt binding to the WRPW motif. The L692F and L651F mutations specifically disrupt gro binding to WRPW but not eh1. The V435A and V435L mutations have no effect on gro binding to either the WRPW motif or eh1 domain.

A large scale genetic screen for enhancers of hairy recovered 50 novel gro alleles, including 7 mutations of the gro WD domain. These mutations, along with additional residues identified in a crystal structure of the human homolog TLE, were assessed for their effects on gro interactions with WRPW and eh1 repressor proteins.

Source was cell extract of S2R+ cell line; proteins produced from transfected construct or endogenous gene.

HGScore = 20.414541