Dimorphic aggregation behavior of a fusion polypeptide incorporating a stable protein domain (EGFP) with an amyloidogenic sequence (retroCspA)

Abstract 

We describe the behavior of a polypeptide consisting of the genetic fusion of a structurally stable single-domain protein, EGFP (an analog of the green fluorescent protein) with an amyloidogenic sequence, retroCspA (known to readily form amyloid fibrils). Refolding of the fusion protein through single-step removal of denaturant and salt results in precipitation into amyloid aggregates displaying fibrillar morphology, thioflavin T binding as well as green fluorescence. Refolding through step-wise reduction of denaturant concentration in the presence of salt yields a soluble aggregate containing a folded, thermally-stable, non-fluorescent EGFP domain. Together, these results indicate that retroCspA forces the fusion protein to aggregate; however, the EGFP domain remains folded in a native-like structural format in both soluble aggregates and precipitates.

Keywords: Beta sheet propagation, Amyloid formation, Stable protein domains, Fusion proteins, Conformational change