Enhanced protein synthesis and secretion using a rational signal-peptide library approach as a tailored tool

Stern, Beate; Optun, Asta; Gey, Christine; Gräfe, Markus; Pryme, Ian F.

doi:10.1186/1753-6561-5-s8-o13

Cited by 6 publications

(8 citation statements)

References 4 publications

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“…Signal peptide derived from human albumin more than doubled the average specific productivity of a stable mAb-producing CHO cell mini pool to almost 40 pg/cell/day compared with using a native IgG LC signal peptide [80]. A signal peptide toolbox has also been generated to identify tailored signal peptides for each protein [81].…”

Section: Vector Design » Signal Peptide and Codon Optimizationmentioning

confidence: 99%

Generation of monoclonal antibody-producing mammalian cell lines

Ho¹,

Yang²

2013

Pharmaceutical Bioprocessing

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Section: Vector Design » Signal Peptide and Codon Optimizationmentioning

confidence: 99%

Generation of monoclonal antibody-producing mammalian cell lines

Ho¹,

Yang²

2013

Pharmaceutical Bioprocessing

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“…The appropriate selection of a SP can have important consequences on protein overexpression, with some authors reporting levels of expression increased by several-fold [82], [83], [84]. Studies have shown that, despite their heterogeneity, many SPs are functionally interchangeable even between different species [85].…”

Section: Focus On the Enhancement Of Translationmentioning

confidence: 99%

Section: Focus On the Enhancement Of Translationmentioning

confidence: 99%

“…As an example, UTR-Tailortech allows the rational design of SP libraries randomized at chosen codon positions [84]. This tool was developed by comparing the success of individual SPs with their amino acid composition and has allowed to predict with respect to which amino acid in which positions can have a decisive influence on protein synthesis or secretion [84]. In contrast to a traditional random approach, which would result in extremely large libraries difficult to manage, libraries generated with UTR-Tailortech are substantially smaller while simultaneously being enriched for good candidates.…”

Section: Focus On the Enhancement Of Translationmentioning

confidence: 99%

“…When combined with high-throughput screening technologies, a tailored SP for any specific protein (including difficult-to-express proteins) can be quickly defined [84].…”

Section: Focus On the Enhancement Of Translationmentioning

confidence: 99%

See 2 more Smart Citations

Engineering Translation in Mammalian Cell Factories to Increase Protein Yield: The Unexpected Use of Long Non-Coding SINEUP RNAs

Zucchelli

Patrucco

Persichetti

et al. 2016

Computational and Structural Biotechnology Journal

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Mammalian cells are an indispensable tool for the production of recombinant proteins in contexts where function depends on post-translational modifications. Among them, Chinese Hamster Ovary (CHO) cells are the primary factories for the production of therapeutic proteins, including monoclonal antibodies (MAbs). To improve expression and stability, several methodologies have been adopted, including methods based on media formulation, selective pressure and cell- or vector engineering. This review presents current approaches aimed at improving mammalian cell factories that are based on the enhancement of translation. Among well-established techniques (codon optimization and improvement of mRNA secondary structure), we describe SINEUPs, a family of antisense long non-coding RNAs that are able to increase translation of partially overlapping protein-coding mRNAs. By exploiting their modular structure, SINEUP molecules can be designed to target virtually any mRNA of interest, and thus to increase the production of secreted proteins. Thus, synthetic SINEUPs represent a new versatile tool to improve the production of secreted proteins in biomanufacturing processes.

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Endoplasmic reticulum‐directed recombinant mRNA displays subcellular localization equal to endogenous mRNA during transient expression in CHO cells

Kallehauge

Kol

Andersen

et al. 2016

Biotechnology Journal

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When expressing pharmaceutical recombinant proteins in mammalian cells, the protein is commonly directed through the secretory pathway, in a signal peptide-dependent manner, to acquire specific post-translational modifications and to facilitate secretion into the culture medium. One key premise for this is the direction of the mRNA encoding the recombinant protein to the surface of the endoplasmic reticulum (ER) for subsequent protein translocation into the secretory pathway. To evaluate the efficiency of this process in Chinese hamster ovary (CHO) cells, the subcellular localization of recombinant mRNA encoding the therapeutic proteins, erythropoietin (EPO) and Rituximab, was determined. The results show that ER-directed recombinant mRNAs exhibited an efficient recruitment to the ER when compared to an endogenous ER-directed mRNA, with no cytoplasmic translation of ER-directed recombinant proteins observed. These observations indicate that the recombinant mRNA, encoding ER-directed proteins, follows the same distribution pattern as endogenous mRNA directed towards the ER. Furthermore, the previous established fractionation method proves to be an efficient tool to study not only recombinant mRNA localization, but also recombinant protein trafficking between the ER and cytosol in CHO cells.

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Enhanced protein synthesis and secretion using a rational signal-peptide library approach as a tailored tool

Cited by 6 publications

References 4 publications

Generation of monoclonal antibody-producing mammalian cell lines

Generation of monoclonal antibody-producing mammalian cell lines

Engineering Translation in Mammalian Cell Factories to Increase Protein Yield: The Unexpected Use of Long Non-Coding SINEUP RNAs

Endoplasmic reticulum‐directed recombinant mRNA displays subcellular localization equal to endogenous mRNA during transient expression in CHO cells

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