Structural constraints on GFP polypeptide insertion. A) GFP trap hotspots: the left chart plots the relative frequency of GFP insertions in introns between exon-exon boundaries comprising predicted structural domains, regions of intrinsic disorder or unclassified regions. The right chart plots un-trapped introns from the same genes and shows a reduction in the intrinsically disordered category. B) Consequences of GFP insertions: in cases where GFP insertions fall into predicted structural domains, mapping of domain sequence to known structures of proteins of the same fold shows that it is mostly surface exposed areas that are affected. In both examples, the overall fold of the GFP target domain is unlikely to be affected by the insertion. Note that the linker residues and the C-terminus of GFP (blue dotted lines) are predicted to be highly flexible. The displayed examples show only one possibility for how the GFP domain is structured relative to the host protein.