Journal of Microbiology and Biotechnology

The Korean Society for Microbiology and Biotechnology (한국미생물·생명공학회)
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In Vitro Formation of Protein Nanoparticle Using Recombinant Human Ferritin H and L Chains Produced from E. coli

  • RO HYEON SU (BioNanotechnology Research Center, Korea Research Institute of Bioscience and Biotechnology) ;
  • PARK HYUN KYU (BioNanotechnology Research Center, Korea Research Institute of Bioscience and Biotechnology) ;
  • KIM MIN GON (BioNanotechnology Research Center, Korea Research Institute of Bioscience and Biotechnology) ;
  • CHUNG BONG HYUN (BioNanotechnology Research Center, Korea Research Institute of Bioscience and Biotechnology)
  • Published : 2005.04.01
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Abstract

We have conducted in vitro reconstitution study of ferritin from its subunits FerH and FerL. For the reconstitution, FerH was produced from an expression vector construct in Escherichia coli and was purified from a heat treated cell extract by using one-step column chromatography. FerL was expressed as inclusion bodies. The denatured form of FerL was obtained by a simple washing step of the inclusion bodies with 3 M urea. The reconstitution experiment was conducted with various molar ratios of urea-denatured FerH and FerL to make the ferritin nanoparticle with a controlled composition of FerH and FerL. SDS-PAGE analysis of the reconstituted ferritins revealed that the reconstitution required the presence of more than 40 molar$\%$ of FerH in the reconstitution mixture. The assembly of the subunits into the ferritin nanoparticle was confmned by the presence of spherical particles with diameter of 10 nm by the atomic force microscopic image. Further analysis of the particles by using a transmission electron microscope revealed that the reconstituted particles exhibited different percentages of population with dense iron core. The reconstituted ferritin nanoparticles made with molar ratios of [FerH]/[FerL]=l00/0 and 60/40 showed that 80 to $90\%$ of the particles were apoferritin, devoid of iron core. On the contrary, all the particles formed with [FerH]/[FerL]=85/ 15 were found to contain the iron core. This suggests that although FerH can uptake iron, a minor portion of FerL, not exceeding $40\%$ at most, is required to deposit iron inside the particle.

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References

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