Investigation of the effect of different culture conditions on recombinant protein production

Year 2023, Volume: 4 Issue: 2, 42 - 48, 31.12.2023

İlkgül Akmayan

https://doi.org/10.51539/biotech.1226205

Cited By: 2

Abstract

After the COVID-19 pandemic, vaccine production technologies have become the focus of attention of researchers. As a matter of fact, recombinant protein-based antigen production, which is one of them, has taken its place in the first place. Proteins obtained by recombinant DNA technology are used in many industrial areas, especially vaccine applications, due to their reliability. Therefore, it is very important to produce targeted recombinant proteins in large quantities. This study, for the high amounts production of Omp25 protein, which is used as a vaccine candidate against brucellosis, in laboratory conditions, is aimed to reveal the effects of conditions that are the pre-culturing process, inoculation in LB or TB media, denatured or native purification, culturing with/without IPTG. All the results were analyzed by SDS-PAGE, confirmed Western Blot, and the total protein amounts were measured Bradford method. According to the results, Omp25 protein could not be obtained under native purification conditions in both cultures without induction, but it was observed under denatured conditions. This result can be explained that the protein in the cell is either misfolded or incorporated into the membrane. The amount of protein appears to be much higher in the presence of the inducer in both media inoculated with the starter pre-culture compared to the overnight pre-culture; 8.79 mg and 39.4 mg from 1 L culture, respectively. Additionally, as expected, the addition of IPTG increased the amount of protein, approximately one-and-a-half-fold for LB and about three-fold for TB. Finally, it was observed that TB medium provided higher protein production than LB, which can be explained by the presence of glycerol and high yeast extract in the medium. Although our study contains results that will attract the attention of vaccine industry, it should be kept in mind that all process should always be optimized depending on the structure of the targeted protein and thus the production amount can be further increased.

Keywords

Recombinant protein, protein amount, culture condition, Omp25

Supporting Institution

Yıldız Technical University Scientific Research Projects Coordination Unit.

Project Number

Project number: FBA-2022-4648.

Thanks

The authors are grateful to the Scientific and Technological Research Council of Turkey (TUBITAK, “2247/C STAR COVID-19”) and The Council of Higher Education of the Republic of Turkey (YOK, “YOK100/2000”) for providing the first author a research scholarship. This report involve a part of İlkgül Akmayan's doctoral thesis. This work was supported by Yıldız Technical University Scientific Research Projects Coordination Unit. Project number: FBA-2022-4648. The authors thank Tuğba ATABEY for her contributions to the rOmp25 protein study.

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