Recombinant-phospholipase A2 production and architecture of inclusion bodies are affected by pH in Escherichia coli

Bacterial inclusion bodies are industrially exploitable amyloids

“…Those IBs formed under uncontrolled increasing alkaline conditions (up to 8.5) have a lower content of amyloid structures and are easier to solubilize using proteinase K or to denature by chaotropic agents if compared with those IBs recovered from bacteria cultured at controlled pH (Castellanos-Mendoza, et al, 2014). Similar results were found when IBs were produced under constant basic pH of 8.5, using different model proteins and genetic backgrounds (Calcines-Cruz, et al, 2018).…”

“…When visualizing the protein production process as a whole, the rate at which the proteins fold in the cell can be reduced by adjusting the growth temperature of the culture, the strength of the gene promoter or even the medium composition. In the first case, the growth rate of bacteria directly affects the metabolism of the cell and specifically protein synthesis (Bosdriesz, et al, 2015, Calcines-Cruz, et al, 2018, Wurm, et al, 2018. In the second scenario, both the transcription and translation rates for the nascent polypeptide are reduced improving the performance of the folding modulators in the same way.…”

“…The environment can also influence IBs formation. In fact, during recombinant protein production, temperature, culture time, nature of the inducer, growth rate, agitation and pH (Table 1, and Supplementary Table 1), have a profound impact on architectonic IB properties such as size, inner structure, content of active protein and their potential for being released or solubilized , Margreiter, et al, 2008, Castellanos-Mendoza, et al, 2014, Krauss, et al, 2017, Valdez-Cruz, et al, 2017, Calcines-Cruz, et al, 2018.…”

“…Even more, IBs formed at 42°C present greater resistance to solubilization and denaturation compared with those formed at 25°C, indicating that the culture temperature determines the conformational properties and stability of the proteins trapped within aggregates (Jevsevar, et al, 2005, de Groot & Ventura, 2006, Vera, et al, 2007. pH has also been examined regarding the formation and quality of IBs (Strandberg & Enfors, 1991, Castellanos-Mendoza, et al, 2014, Calcines-Cruz, et al, 2018. When the pH dropped below 5.5, an increase in the relative amount of IBs was observed compared with cultures developed at constant pH, upon temperature-mediated induction of recombinant gene expression (Strandberg & Enfors, 1991).…”

Soluble recombinant pyruvate oxidase production in Escherichia coli can be enhanced and inclusion bodies minimized by avoiding pH stress

“…These results are consistent with those on fusion proteins of Staphylococcus aureus protein A and E. coli β ‐galactosidase and blue fluorescent protein . Several researchers have investigated the effects of pH on the morphology of inclusion bodies, and the results show a significant influence for pH on inclusion body formation . A high pH can increase inclusion body formation in E. coli , although changes in soluble heterologous proteins were are not always evident .…”