Overexpression in Escherichia coli of the AT-rich trpA and trpB genes from the hyperthermophilic archaeon Pyrococcus furiosus.
Expression of AT-rich genes from microorganisms such as archaea is often inefficient in Escherichia coli. The trpA and trpB genes encoding the tryptophan synthase subunits were cloned from the hyperthermophilic archaeon Pyrococcus furiosus. No apparent difference in codon bias was found between the genes. However, using a conventional cloning vector having the lac promoter, the trpB gene was expressed poorly in E. coli, whereas the trpA gene was overexpressed. The expression of the trpB gene was remarkably enhanced (>12-fold) by the introduction of an overlapping leader open reading frame. The expression of the trpA gene was also improved ( approximately 1.5-fold). This approach may be useful for overexpressing various kinds of AT-rich genes.
Ishida
Masami
M
Laboratory of Marine Biochemistry, Tokyo University of Fisheries, Konan 4, Minato-ku, Tokyo 108-8477, Japan. ishida@tokyo-u-fish.ac.jp
Oshima
Tairo
T
Yutani
Katsuhide
K
D020856:AT Rich Sequence / Q000235:genetics*; D019843:Archaeal Proteins / Q000096:biosynthesis* / Q000235:genetics; D001483:Base Sequence; D003001:Cloning, Molecular; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics; D015870:Gene Expression; D005796:Genes; D005822:Genetic Vectors / Q000235:genetics; D008969:Molecular Sequence Data; D016296:Mutagenesis; D009247:NADH, NADPH Oxidoreductases / Q000096:biosynthesis* / Q000235:genetics; D019711:Pyrococcus / Q000201:enzymology / Q000235:genetics*; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D014364:Tryptophan / Q000096:biosynthesis
D019843:Archaeal Proteins; D011994:Recombinant Proteins; D014364:Tryptophan; D009247:NADH, NADPH Oxidoreductases; C049608:trypanothione reductase