High level expression in Escherichia coli of a fungal gene under the control of strong promoters.
A recent report (Patino et al., (1989) FEMS Microbiol. Lett. 58, 139-144) described the low level expression, in Escherichia coli, of the Isopenicillin N Synthase (IPNS) gene from Cephalosporium acremonium under the control of strong promoters. We report here our work on the expression of the IPNS gene. Plasmids containing the IPNS gene under the control of the trp or trc promoters directed synthesis of high levels of active IPNS in E. coli. Constitutive and inductive high level IPNS expression systems have been developed. Importantly, the expression vectors do not encode beta-lactamase so IPNS activity can be determined directly by biological assays. Analysis by nmr verified that the IPNS produced from these expression systems catalysed the conversion of delta-(L-alpha-aminoadipoyl)-L-cysteinyl-D-valine (LLD-ACV) to isopenicillin N in high yield.
Baldwin
J E
JE
Dyson Perrins Laboratory, Oxford, U.K.
Blackburn
J M
JM
Schofield
C J
CJ
Sutherland
J D
JD
D000164:Acremonium / Q000201:enzymology / Q000235:genetics*; D001483:Base Sequence; D004271:DNA, Fungal; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics*; D015966:Gene Expression Regulation, Fungal / Q000235:genetics*; D005800:Genes, Fungal; D005822:Genetic Vectors; D008969:Molecular Sequence Data; D010088:Oxidoreductases / Q000096:biosynthesis / Q000235:genetics*; D010957:Plasmids; D011401:Promoter Regions, Genetic
D004271:DNA, Fungal; D010088:Oxidoreductases; C038370:isopenicillin N synthetase