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Nucleic Acids Research, 1993, Vol. 21, No. 18 4268-4271
© 1993

MOLECULAR BIOLOGY

Influence of the three nucleotides upstream of the initiation codon on expression of the Escherichia coli lacz gene in Saccharomyces Cerevisiae

Alfred C. Looman* and Jan Albert Kuivenhoven1,+

Institut fuür Gaürungsgewerbe and Biotechnologie Seestraße 13, D 13353 Berlin 1Technische Universitaü Berlin, Abt, Mikrobiologie and Genetik Gustave-Meyer-Alle 25, D 13355 Berlin, Germany

*To whom correspondence should be addressed at: Technische Universitit Berlin, Abt. Microbiologie and Genetik, Gustav-Meyer-Allee 25, D 13355 Berlin, Germany

Received May 28, 1993. Revised August 4, 1993. Accepted August 4, 1993.

By introducing synthetic oligonucleotldes into a lacZyeast expression vector a set of 47 plasmlda (out of 64 possible) was generated, differing only In the three bases immediately upstream of the AUG Initiation codon of the Escherichia coll lacZ gene. Expression of the jS-galactosldase fusion protein encoded by the different plasmids was determined In Saccharomyces cerevisiae by Immunogel electrophoresls. Among the clones tested we found a factor 3 difference in expression. A slight nucleotlde preference was found In positions -3(A<G<C = U) and - 2 ( G < C = U<A). The choice of the nucleotide at position -1 immediately 5' of the AUG did not effect translation efficiency. Increasing homology to the yeast consensus sequence (AAAAAAAUGUCU) was not concomitant with an Increased translation efficiency. Our results Indicate that the choice of nucleotides Immediately preceding the initiation codon in yeast does not dramatically Influence translation efficiency, as in prokaryotes or higher eukaryotes.

+Present address: Academic Medical Center, Lipid Research Group, Meilbergdreef 9, NL 1105 AZ, Amsterdam, The Netherlands


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