Optimisation of protein synthesis rates in reticulocyte lysates and reconstituted systems |
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| Authors: | N Desai K L Manchester |
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| Affiliation: | 1. Inflammation Research Center, VIB, Ghent, Belgium;2. Department for Biomedical Molecular Biology, University Ghent, Ghent, Belgium;1. Department of Biology and Biological Engineering, Chalmers University of Technology, SE41296 Gothenburg, Sweden;2. Novo Nordisk Foundation Center for Biosustainability, Chalmers University of Technology, SE41296 Gothenburg, Sweden;3. Novo Nordisk Foundation Center for Biosustainability, Technical University of Denmark, DK2970 Hørsholm, Denmark;1. Department of Life Sciences and the National Institute for Biotechnology in the Negev, Ben-Gurion University of the Negev, Be’er Sheva 84105, Israel;1. Department of Chemistry and Biochemistry, New Brunswick Centre for Precision Medicine, Université de Moncton, Moncton, New Brunswick, Canada;2. Dr. George-L.-Dumont University Hospital Centre, Moncton, New Brunswick, Canada;1. Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, MA 02139, United States;2. Department of Chemical and Biomolecular Engineering, Tulane University, New Orleans, LA 70118, United States |
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| Abstract: | Means of increasing the very low activity of a reconstituted protein synthesising system from rabbit reticulocytes were investigated. Increasing the concentration of labelled amino acid, addition of polyamines, use of Sepharose-filtered as opposed to centrifuged ribosomes, use of untreated as opposed to gel-filtered cytosol and an increase in ratio of cytosol to ribosomes all contributed to the increase in activity of the system to the point where activity was clearly consistent with initiation taking place. Similar activities could not be attained with rat liver cytosol though rat liver ribosomes incorporated well in reticulocyte cytosol. Incorporation by lysates was also found to be dependent on the concentration of the labelled amino acid added. |
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