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Upregulation of YciM Expression Reduces Endotoxin Contamination of Recombinant Proteins Produced in Escherichia coli Cells

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Abstract

Recombinant proteins produced in Escherichia coli are often contaminated with endotoxins, which can be a serious problem for their further application. One of the possible solutions is the use of modified strains with reduced lipopolysaccharide (LPS) levels. We compared two approaches to engineering such strains. The first commonly known approach was modification of LPS biosynthesis pathway by knocking out seven genes in the E. coli genome. The second approach, which has not been previously used, was to increase expression of E. coli protein YciM. According to the published data, elevated expression of YciM leads to the reduction in the amount of the LpxC enzyme involved in LPS biosynthesis. We investigated the impact of YciM coexpression with eGFP on the content of endotoxins in the purified recombinant eGFP samples. Both approaches provided similar outcomes, i.e., decreased the endotoxin levels in the purified protein samples.

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Abbreviations

LPS:

lipopolysaccharide

OD:

optical density

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Funding

This research was supported by the Russian Science Foundation (project no. 23-24-00080, https://rscf.ru/project/23-24-00080/ [in Russian]).

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Authors and Affiliations

  1. Lopukhin Federal Research and Clinical Center of Physical-Chemical Medicine of Federal Medical Biological Agency, 119435, Moscow, Russia

    Pavel A. Bobrovsky, Daria D. Kharlampieva, Sergey A. Kirillin, Ksenia A. Brovina, Ekaterina N. Grafskaia, Vassili N. Lazarev & Valentin A. Manuvera

  2. Moscow Institute of Physics and Technology (National Research University), 141701, Moscow, Russia

    Pavel A. Bobrovsky, Ksenia A. Brovina, Vassili N. Lazarev & Valentin A. Manuvera

Authors
  1. Pavel A. Bobrovsky
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  2. Daria D. Kharlampieva
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  3. Sergey A. Kirillin
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  4. Ksenia A. Brovina
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  5. Ekaterina N. Grafskaia
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  6. Vassili N. Lazarev
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  7. Valentin A. Manuvera
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Contributions

P.A.B. and S.A.K. conducted genetic engineering experiments; D.D.Kh. performed bacterial endotoxin assays; K.A.B. and E.N.G. conducted microbiological experiments; V.A.M. purified the recombinant protein; V.N.L. supervised the study; P.A.B., D.D.Kh., and V.A.M. wrote the manuscript.

Corresponding author

Correspondence to Valentin A. Manuvera.

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The authors declare no conflict of interest. This manuscript does not contain description of studies involving humans or animals performed by any of the authors.

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Bobrovsky, P.A., Kharlampieva, D.D., Kirillin, S.A. et al. Upregulation of YciM Expression Reduces Endotoxin Contamination of Recombinant Proteins Produced in Escherichia coli Cells. Biochemistry Moscow 88, 1318–1325 (2023). https://doi.org/10.1134/S0006297923090110

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  • DOI: https://doi.org/10.1134/S0006297923090110

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