Optimization of recombinant membrane protein production in the engineered Escherichia coli strains SuptoxD and SuptoxR

Myrsini Michou, Charalampos Kapsalis, Christos Pliotas, Georgios Skretas

Research output: Contribution to journalArticlepeer-review

19 Citations (Scopus)
8 Downloads (Pure)

Abstract

Membrane proteins (MPs) execute a wide variety of critical biological functions in all living organisms and constitute approximately half of current targets for drug discovery. As in the case of soluble proteins, the bacterium Escherichia coli has served as a very popular overexpression host for biochemical/structural studies of membrane proteins as well. Bacterial recombinant membrane protein production, however, is typically hampered by poor cellular accumulation and severe toxicity for the host, which leads to low levels of final biomass and minute volumetric yields. In previous work, we generated the engineered E. coli strains SuptoxD and SuptoxR, which upon coexpression of the effector genes djlA or rraA, respectively, can suppress the cytotoxicity caused by MP overexpression and produce enhanced MP yields. Here, we systematically looked for gene overexpression and culturing conditions that maximize the accumulation of membrane-integrated and well-folded recombinant MPs in these strains. We have found that, under optimal conditions, SuptoxD and SuptoxR achieve greatly enhanced recombinant production for a variety of MP, irrespective of their archaeal, eubacterial, or eukaryotic origin. Furthermore, we demonstrate that the use of these engineered strains enables the production of well-folded recombinant MPs of high quality and at high yields, which are suitable for functional and structural studies. We anticipate that SuptoxD and SuptoxR will become broadly utilized expression hosts for recombinant MP production in bacteria.
Original languageEnglish
JournalACS Synthetic Biology
VolumeArticles ASAP
Early online date6 Jun 2019
DOIs
Publication statusE-pub ahead of print - 6 Jun 2019

Keywords

  • Recombinant membrane protein production
  • Toxicity
  • E. coli SuptoxD
  • E. coli SuptoxR
  • DjLA
  • RraA

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