{"id":"doi:10.1371/journal.pone.0082234","kind":"article","title":"IRES-mediated translation of membrane proteins and glycoproteins in eukaryotic cell-free systems","authors":["Brödel AK","Sonnabend A","Roberts LO","Stech M","Wüstenhagen DA","Kubick S"],"author_count":6,"year":2013,"published":"2013-12-20","venue":"PloS one","cited_by":78,"doi":"10.1371/journal.pone.0082234","arxiv_id":null,"nct_id":null,"source_url":"https://doi.org/10.1371/journal.pone.0082234","archive_url":"https://space-kitty.com/papers/doi:10.1371/journal.pone.0082234","updated":null,"pmid":"24376523","pmcid":"PMC3869664","openalex_id":null,"pdf_url":"https://europepmc.org/articles/PMC3869664/pdf","abstract":"Internal ribosome entry site (IRES) elements found in the 5' untranslated region of mRNAs enable translation initiation in a cap-independent manner, thereby representing an alternative to cap-dependent translation in cell-free protein expression systems. However, IRES function is largely species-dependent so their utility in cell-free systems from different species is rather limited. A promising approach to overcome these limitations would be the use of IRESs that are able to recruit components of the translation initiation apparatus from diverse origins. Here, we present a solution to this technical problem and describe the ability of a number of viral IRESs to direct efficient protein expression in different eukaryotic cell-free expression systems. The IRES from the intergenic region (IGR) of the Cricket paralysis virus (CrPV) genome was shown to function efficiently in four different cell-free systems based on lysates derived from cultured Sf21, CHO and K562 cells as well as wheat germ. Our results suggest that the CrPV IGR IRES-based expression vector is universally applicable for a broad range of eukaryotic cell lysates. Sf21, CHO and K562 cell-free expression systems are particularly promising platforms for the production of glycoproteins and membrane proteins since they contain endogenous microsomes that facilitate the incorporation of membrane-spanning proteins and the formation of post-translational modifications. We demonstrate the use of the CrPV IGR IRES-based expression vector for the enhanced synthesis of various target proteins including the glycoprotein erythropoietin and the membrane proteins heparin-binding EGF-like growth factor receptor as well as epidermal growth factor receptor in the above mentioned eukaryotic cell-free systems. CrPV IGR IRES-mediated translation will facilitate the development of novel eukaryotic cell-free expression platforms as well as the high-yield synthesis of desired proteins in already established systems.","abstract_license":"CC-BY (article license as recorded by Europe PMC; abstract reproduced unmodified apart from markup removal)","license":"CC-BY","is_open_access":true,"retracted":false,"peer_reviewed":null,"categories":[],"details":null,"provenance":"europepmc","attribution":"Abstract of \"IRES-mediated translation of membrane proteins and glycoproteins in eukaryotic cell-free systems\" by Brödel AK, Sonnabend A, Roberts LO et al., https://doi.org/10.1371/journal.pone.0082234, used under its Creative Commons Attribution license (CC-BY); markup removed, otherwise unmodified.","topics":[{"slug":"synthetic-biology","name":"Synthetic Biology & Genome Engineering","domain_slug":"biotech"}],"cited_in_dossiers":[]}