✍ Scribed by Joop J. Heuvel; Hendrik A. Raué
No coin nor oath required. For personal study only.
Synthetic oligonucleotides encoding the 5'non-translated (leader) sequence of the coat protein mRNA of alfalfa mosaic virus RNA 4 or the f~' leader sequence of tobacco mosaic virus RNA were used to replace the natural leader region of the yeast phosphoglycerate kinase (PGK1) mRNAs and the translational efficiency of the chimeric mRNA was determined in yeast cells. In neither case did we observed a significant increase compared to the translational efficiency shown by the wild-type P G K mRNA, in contrast to the known stimulatory effect of these leader sequences on translation in mammalian, plant and bacterial in-vivo a n d / o r in-vitro systems. The same result was obtained when the translational efficiencies in yeast cells of Escherichia coli fl-galactosidase mRNAs carrying the P G K or either of the two viral leader sequences were compared.
As part of our efforts to optimize biotechnological production of (heterologous) proteins by yeast cells we have recently investigated the effect of systematic changes in either the length or nucleotide composition of the leader sequence on translational efficiency in Saccharomyces cerevisiae, using the yeast phosphoglycerate kinase gene (PGK1) m R N A as a model (van den Heuvel et al. 1989(van den Heuvel et al. , 1990)). While these studies provided valuable information concerning the relationship between the rate of translation and these two aspects of leader structure, none of the changes resulted in significant improvement in translational efficiency. We therefore decided to investigate the possible usefulness of the AMV RNA 4 a n d / o r TMV g)' leader sequence as a tool for increasing the efficacy of biotechnological production of (heterologous) proteins in yeast cells.
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