A protein whose main function is to bring other
proteins together for them to interact. These
proteins usually have many protein binding domains
(like WD40 repeats). Below are some experiments
that can be done to see if a protein acts as a
scaffold in vivo:
- Defined interactions between interacting
proteins (yeast 2-hybrid)
- IP (or GST pull downs) scaffold protein. Do
other proteins come down with the scaffold?
- Is scaffold protein needed? What happens in
a scaffold protein null?
- Use GST fused to different scaffold domains
to determine modular organization of scaffold
- Does loss of scaffold protein result in mislocalization
of other proteins? (immunofluorescences)
- Does loss of scaffold protein result in instability
of target proteins: (immunoblot)
- Does binding one target rely on binding to
others? Look at IP in different muant backgrounds.