Tryptophan promoter derivatives on multicopy plasmids: a comparative analysis of expression potentials in Escherichia coli.
A collection of variant plasmids expressing either Escherichia coli galactokinase or human serum albumin under the control of several E. coli trp promoter derivatives were constructed and studied for both efficiency of expression and regulation by tryptophan. Several variables, including the length of the upstream region, tandem duplications of a core promoter, and the insertion of the trp repressor trpR gene onto the expression vector, were studied. It is shown that derivatives containing sequences upstream from the -35 region or multiple copies of the trp promoter produce twofold higher levels of protein than plasmids with a minimal trp promoter truncated at -40. We show that the expression of a heterologous protein such as albumin can be significantly improved (13% vs. 7% of total proteins) if both the upstream trp promoter region, which enhances promoter strength, and an intact trpR gene, are included on the plasmids.
Laboratoire de Génétique, Institut de Biotechnologie, Rhône-Poulenc Santé, France.
M
M
Latta
M
M
Philit
I
I
Maury
F
F
Soubrier
P
P
Denèfle
J F
JF
Mayaux
D001483:Base Sequence; D004926:Escherichia coli / Q000235:genetics*; D005686:Galactokinase / Q000096:biosynthesis; D015964:Gene Expression Regulation, Bacterial; D005822:Genetic Vectors; D006801:Humans; D008969:Molecular Sequence Data; D010957:Plasmids; D011401:Promoter Regions, Genetic / Q000235:genetics*; D011994:Recombinant Proteins / Q000096:biosynthesis; D012097:Repressor Proteins / Q000235:genetics; D012709:Serum Albumin / Q000096:biosynthesis; D014158:Transcription, Genetic / Q000235:genetics; D014364:Tryptophan / Q000235:genetics*
D011994:Recombinant Proteins; D012097:Repressor Proteins; D012709:Serum Albumin; D014364:Tryptophan; D005686:Galactokinase