Summary
All five tryptophan biosynthetic genes of Saccharomyces cerevisiae were unified on plasmid pME554, which is based on 2 μm DNA and pBR322 sequences allowing for autonomous replication in yeast and E. coli. Homologous and heterologous expression of this artificial yeast TRP-gene cluster was studied. Plasmid pME554 allowed for nearly normal growth of a yeast strain bearing auxotrophic mutations in all five TRP-genes. The plasmid-borne genes TRP2 to TRP5 were expressed and regulated normally in the frame of the general control. Gene TRP1, carried on an EcoRI/BglII fragment lacking the ARS1 function, was expressed poorly and did not respond to the general control like the chromosomally-borne TRP1 gene.
Plasmid pME554 allowed for poor growth of E. coli strain W3110 tna − ΔtrpEA2 on minimal medium. Marked stimulation was observed, however, when anthranilic acid or indole were added. Accordingly, poor expression of the first Trp-enzyme anthranilate synthase and the last enzyme tryptophan synthase was found, whereas the other three genes were moderately well expressed in E. coli.
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Abbreviations
- bp:
-
basepair
- kb:
-
kilobase
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Niederberger, P., Aebi, M., Furter, R. et al. Expression of an artificial yeast TRP-gene cluster in yeast and Escherichia coli . Mol Gen Genet 195, 481–486 (1984). https://doi.org/10.1007/BF00341450
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DOI: https://doi.org/10.1007/BF00341450