| Autor: |
Benjamin Strutton, Stephen RP Jaffe, Caroline A Evans, Gregory JS Fowler, Paul D Dobson, Jagroop Pandhal, Phillip C Wright |
| Jazyk: |
angličtina |
| Rok vydání: |
2019 |
| Předmět: |
|
| Zdroj: |
Bioengineering, Vol 6, Iss 1, p 27 (2019) |
| Druh dokumentu: |
article |
| ISSN: |
2306-5354 |
| DOI: |
10.3390/bioengineering6010027 |
| Popis: |
Escherichia coli strains have been modified in a variety of ways to enhance the production of different recombinant proteins, targeting membrane protein expression, proteins with disulphide bonds, and more recently, proteins which require N-linked glycosylation. The addition of glycans to proteins remains a relatively inefficient process and here we aimed to combine genetic modifications within central carbon metabolic pathways in order to increase glycan precursor pools, prior to transfer onto polypeptide backbones. Using a lectin screen that detects cell surface representation of glycans, together with Western blot analyses using an O-antigen ligase mutant strain, the enhanced uptake and phosphorylation of sugars (ptsA) from the media combined with conservation of carbon through the glyoxylate shunt (icl) improved glycosylation efficiency of a bacterial protein AcrA by 69% and over 100% in an engineered human protein IFN-α2b. Unexpectedly, overexpression of a gene involved in the production of DXP from pyruvate (dxs), which was previously seen to have a positive impact on glycosylation, was detrimental to process efficiency and the possible reasons for this are discussed. |
| Databáze: |
Directory of Open Access Journals |
| Externí odkaz: |
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