Abstract
We report on the construction of functionalized nanotubes based on tail sheath protein 041 from vB_KleM-RaK2 bacteriophage. The truncated 041 protein (041Δ200) was fused with fluorescent proteins GFP and mCherry or amidohydrolase YqfB. The generated chimeric proteins were successfully synthesized in E. coli BL21 (DE3) cells and self-assembled into tubular structures. We detected the fluorescence of the structures, which was confirmed by stimulated emission depletion microscopy. When 041Δ200GFP and 041Δ200mCherry were coexpressed in E. coli BL21 (DE3) cells, the formed nanotubes generated Förster resonance energy transfer, indicating that both fluorescent proteins assemble into a single nanotube. Chimeric 041Δ200YqfB nanotubes possessed an enzymatic activity, which was confirmed by hydrolysis of N4-acetyl-2′-deoxycytidine. The enzymatic properties of 041Δ200YqfB were similar to those of a free wild-type YqfB. Hence, we conclude that 041-based chimeric nanotubes have the potential for the development of delivery vehicles and targeted imaging and are applicable as scaffolds for biocatalysts.
Highlights
Nanoscale materials based on self-assembly of proteins are used for various purposes, including the formation of structurally different shapes; the development of biosensors; the manufacture of optical, conductive, semiconductor, and magnetic nanoelectronics materials; as well as in gene and drug-delivery devices and vaccines [1,2,3]
Coomassie Plus Assay Reagent were purchased from Thermo Fisher Scientific, Vilnius, Lithuania. pET21d and pET28b vectors were purchased from Novagen, Madison, WI, USA
We investigated the stability of fluorescent nanotubes by incubating the purified nanotubes at different temperatures or pH for 1 h and by analysis using size exclusion chromatography (SEC)
Summary
Nanoscale materials based on self-assembly of proteins are used for various purposes, including the formation of structurally different shapes; the development of biosensors; the manufacture of optical, conductive, semiconductor, and magnetic nanoelectronics materials; as well as in gene and drug-delivery devices and vaccines [1,2,3]. As a natural source of countless self-assembling proteins, are widely studied and adapted for these purposes [4,5]. They come in a wide variety of shapes and sizes. A chemical-modification strategy has been used for the immobilization of enzymes through streptavidin-biotin assembly or glutaraldehyde [6,7]. These methods have some disadvantages, such as multistep and low efficiency. The development and testing of novel nanomaterials are crucial for further progress in this area
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