| dc.description.abstract | Protein-based materials can combine impressive mechanical properties and biocompatibility with a variety of functions through protein fusions, which make the materials useful in a diverse range of applications. Drosophila Ultrabithorax (Ubx) is an intrinsically disordered transcription factor which self-assembles into a variety of materials in vitro. The materials are biocompatible and capable of functionalization with proteins of interest, which retain their function while fused to materials. These properties make Ubx materials ideal for use in many fields.
Here I show that Ubx materials can reversibly, noncovalently bind and protect DNAs harboring the Ubx target DNA sequence. Ubx fibers bind DNA sequence specifically, and DNA with multiple binding sites has a slower rate of release from fibers. Ubx fibers have sustained release of DNA capable of transforming cells for up to 60 days.
Ubx materials can also be functionalized with other proteins, such as a biosensing protein. One proof-of-concept sensor system that has been developed is a set of truncated circular permutants of GFP, collectively known as leave-one-out GFP (LOOn-GFP), in which the nth β-strand has been ‘left out’ of the protein. Without this β-strand the GFP is unable to fold properly and is not fluorescent. Rebinding of peptide leads to recovery of fluorescence, making this construct a self-reporting biosensor for the missing piece of its sequence. However, LOO-GFPs have a tendency to oligomerize, which hampers ligand binding and leads to fluorescence in the unbound state. Ubx offers a transparent support system for LOO-GFPs and has the potential to maintain LOO-GFP stability and binding affinity while preventing LOO-GFP aggregation. Indeed, I found that immobilization on Ubx materials stabilizes LOO-GFP without negatively impacting the chromophore and improves the maximum relative fluorescent intensity of the protein.
This data shows that Ubx materials have promise to be used someday as a vehicle for gene therapy. Ubx materials are particularly suited for use as a DNA delivery scaffold that releases nucleic acids over a long period of time. LOO-GFP-Ubx materials are also promising as biosensors for point of care medical testing or to monitor zoonotic mosquito-borne viruses like dengue fever virus and zika virus. | |
