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The Screening of Refolding Conditions and Obtainment of the Recombinant Antistaphylococcal Endolysin LysKCA in Active Form from E. coli Inclusion Bodies

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Abstract

Step-by-step screening of the main characteristics of refolding buffer is proposed to obtain recombinant antistaphylococcal endolysin LysK containing two catalytic domains, CHAP and amidase-2, in active form from E. coli inclusion bodies. The optimal pH, temperature, redox potential of the refolding buffer, as well as the optimal protein concentration and type of antiaggregation compound were determined. The composition of a renaturation system of antistaphylococcal endolysin containing 20 mM phosphate buffer, pH 7.4, 0.4 M sucrose, and 2.5 mM DTT at 10°C with dilution to a final concentration of ~150 μg/mL was found to be optimal. The refolding yield after scaling was about 29.5 ± 6.7%, which produced 16 ± 2.3 mg of the target protein from 2.25 g of washed inclusion bodies with a specific enzyme activity of 1.8 ± 0.73 × 103 U/mg.

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Authors and Affiliations

  1. Belarusian State University, Biology Department, 220030, Minsk, Belarus

    A. V. Žydziecki, S. G. Golenchenko, U. A. Prakulevich & M. V. Sholukh

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  1. A. V. Žydziecki
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  2. S. G. Golenchenko
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  3. U. A. Prakulevich
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  4. M. V. Sholukh
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Correspondence to A. V. Žydziecki.

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The authors declare that they have no conflict of interest. This article does not contain any studies involving animals or human participants performed by any of the authors.

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Translated by P. Kuchina

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Žydziecki, A.V., Golenchenko, S.G., Prakulevich, U.A. et al. The Screening of Refolding Conditions and Obtainment of the Recombinant Antistaphylococcal Endolysin LysKCA in Active Form from E. coli Inclusion Bodies. Appl Biochem Microbiol 56, 44–50 (2020). https://doi.org/10.1134/S0003683820010160

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