Effect of an amyloidogenic sequence attached to yellow fluorescent protein.

Green fluorescent protein (GFP) is often misfolded into nonfluorescent states when an aggregatable sequence is attached to its N-terminus. However, GFP fusions with highly aggregatable, prion-determining, and highly charged sequences from yeast prions, such as Sup35 and Ure2p, form green fibrils with properly folded GFP. To gain further insight into the general effect of an aggregatable sequence attached to fluorescent protein, we designed eight fusion proteins of a yellow variant of GFP (YFP) containing an aggregation-prone amyloidogenic sequence derived from human medin, attached via different lengths of linker sequence. Seven fusion proteins formed white fibrils lacking native YFP function. However, the fusion with an 18-residue medin sequence and a 50 amino acid linker formed fibrils with yellow color of folded YFP. Deconvolution analysis of infrared spectra also supports the presence of properly folded YFP in the fibrils formed by this protein. These results suggest that, the presence of an amyloidogenic sequence to a folded protein can promote the formation of fibrils and disrupt the native structures whereas the structure of the folded region is retained by optimizing sequences of amyloidogenic and linker regions.