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High-efficient renaturation of immobilized recombinant C-terminal fragment of human alpha-fetoprotein

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Abstract

C-terminal fragment of a human oncofetal alpha-fetoprotein (AFP) may be used in targeted cytostatics delivery to malignant cells of many tumors. AFP fragment (from 404 to 595 amino acids residues of a full-sized protein) was cloned and produced in Escherichia coli cells, BL21 strain (DE3) in the form of inclusion bodies. To obtain a functionally active protein, is it necessary to renature the protein. The renaturation procedure of the AFP third domain (rAFP3D) is considerably complicated by the fact that the protein is hydrophobic and contains a large number of S-S bonds. A renaturation technique of rAFP3D immobilized on silicic metal chelate resin has been developed. The yield of renatured C-terminal fragment was no less than 60% with purity on the order of 98%. The developed technique has been applied for the first time for hydrophobic protein with a large number of S-S bonds. The approach can be applied for efficient renaturation of other hydrophobic proteins with a large number of disulfide bonds for scientific and practical purposes.

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Abbreviations

AFP:

human alpha-fetoprotein

AFPR:

receptor for AFP

IB:

inclusion bodies

rAFP3D:

recombinant AFP third domain

LB:

Luria Bertani medium

PBS:

phosphate buffered saline

CD:

circular dichroism

US:

ultrasound

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Authors and Affiliations

  1. Moscow Research Institute of Medical Ecology, Moscow, 117638, Russia

    O. A. Sharapova & S. E. Severin

  2. All-Russia Center for Molecular Diagnostics and Treatment, Moscow, 117638, Russia

    M. S. Yurkova, S. M. Andronova, A. N. Fedorov & E. S. Severin

Authors
  1. O. A. Sharapova
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  2. M. S. Yurkova
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  3. S. M. Andronova
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  4. A. N. Fedorov
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  5. S. E. Severin
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  6. E. S. Severin
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Corresponding author

Correspondence to O. A. Sharapova.

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Original Russian Text © O.A. Sharapova, M.S. Yurkova, S.M. Andronova, A.N. Fedorov, S.E. Severin, E.S. Severin, 2011, published in Prikladnaya Biokhimiya i Mikrobiologiya, 2011, Vol. 47, No. 5, pp. 523–529.

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Sharapova, O.A., Yurkova, M.S., Andronova, S.M. et al. High-efficient renaturation of immobilized recombinant C-terminal fragment of human alpha-fetoprotein. Appl Biochem Microbiol 47, 474–479 (2011). https://doi.org/10.1134/S0003683811050127

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  • DOI: https://doi.org/10.1134/S0003683811050127

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