Expression of recombinant swine ferritin heavy chain with enhanced solubility in Escherichia coli and simplified purification of ferritin nanoparticles.

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Tác giả: Kyoung-Ho Lee, Hyun-Jin Shin, M S B W T M Nipuna Sudaraka Tennakoon

Ngôn ngữ: eng

Ký hiệu phân loại: 809.008 History and description with respect to kinds of persons

Thông tin xuất bản: United States : Protein expression and purification , 2025

Mô tả vật lý:

Bộ sưu tập: NCBI

ID: 708446

 Ferritin is a versatile biomolecule used in various medical applications such as drug delivery, vaccines, biological imaging, and diagnostics. The purity and concentration of the ferritin nanoparticles are crucial for achieving excellent outcomes. In this study, we expressed and purified the recombinant swine ferritin heavy chain (rsFTH) as a new candidate for recombinant ferritin nanoparticles. We generated two types of plasmids that can express rsFTH in mammalian and prokaryotic systems. The myc-tagged rsFTH expressed in the mammalian system was purified and ferritin nanoparticles were validated using dynamic light scattering (DLS) and transmission electron microscopy (TEM). A prokaryotic expression system was used to produce rsFTH on a large scale. Protein expression was optimized in Escherichia coli BL21 under varying temperatures and IPTG conditions, and solubility was enhanced by incubation at 25 °C for 18-22 h in auto-induction media, resulting in approximately >
 50 % protein content in the soluble fraction compared with the pellet. Protein purification was achieved using His-tag affinity chromatography and dialysis with Tris-HCl buffer, yielding adequately pure rsFTH without any apparent protein aggregates. SDS-PAGE and Western blot analysis confirmed the expected molecular weight of rsFTH, and Native-PAGE demonstrated polymerization into higher molecular weight forms. Particle size analysis of purified rsFTH revealed a mean diameter of 15.5 nm, with transmission electron microscopy (TEM) imaging confirming spherical ferritin particles with an iron core. These results suggest that rsFTH can be efficiently expressed and purified in both mammalian and bacterial systems, and has potential applications in nanotechnology and biotechnology.
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