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Abstract <p>The relationship between the key conditions for induction of gene expression in cells of <i>E. coli</i> (DE3) strains in the pET vector system and the functional characteristics of the target protein in whole resting cells is shown on the example of heterologous expression of the <i>kstD2</i> gene encoding 3-ketosteroid-1-dehydrogenase KstD2 from <i>Nocardioides simplex</i> VKM Ac-2033D. The highest activity of 1-dehydrogenation of hydrocortisone to prednisolone was observed in <i>E. coli</i> BL21(DE3) Rosetta/pET_kstD2. The quantity and stability of 3-ketosteroid-1-dehydrogenase activity of the recombinant cells did not correlate with the observed protein accumulation. The maximum yield of the biotransformation product prednisolone increased when using cells induced in the later segment of the logarithmic growth phase, as well as with the shortest duration of induction (6 h). The research results may be in demand in the construction of effective strains producing steroid 1-dehydroanalogues for the pharmaceutical industry.</p>

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Key Factors Influencing the Activity of Actinobacterial 3-Ketosteroid-1-dehydrogenase in E. coli Cells

  • 期刊论文

  • 发表日期：

    2026年5月25日

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  • S. R. Fufaeva,
  • D. V. Dovbnya,
  • A. A. Shutov,
  • M. V. Donova

摘要

Abstract

The relationship between the key conditions for induction of gene expression in cells of E. coli (DE3) strains in the pET vector system and the functional characteristics of the target protein in whole resting cells is shown on the example of heterologous expression of the kstD2 gene encoding 3-ketosteroid-1-dehydrogenase KstD2 from Nocardioides simplex VKM Ac-2033D. The highest activity of 1-dehydrogenation of hydrocortisone to prednisolone was observed in E. coli BL21(DE3) Rosetta/pET_kstD2. The quantity and stability of 3-ketosteroid-1-dehydrogenase activity of the recombinant cells did not correlate with the observed protein accumulation. The maximum yield of the biotransformation product prednisolone increased when using cells induced in the later segment of the logarithmic growth phase, as well as with the shortest duration of induction (6 h). The research results may be in demand in the construction of effective strains producing steroid 1-dehydroanalogues for the pharmaceutical industry.