Development of a prokaryotic expression vector that exploits dicistronic gene organization.
For unknown reasons, levels of expression of foreign genes inserted into expression vectors in Escherichia coli have frequently been undetectable. The most critical step in the successful production of foreign proteins seems to be the initiation of translation. Since most prokaryotic genes are transcribed in a polycistronic form, we have devised a new prokaryotic expression system utilizing dicistronic gene organization. Downstream from a strong promoter and the gene encoding glutathione S-transferase from Schistosoma japonicum, various foreign genes were connected via a ribosome-binding site, a stop codon and a start codon. The VH domain of an immunoglobulin fused to the alpha subunit of tryptophan synthase, FK506-binding protein, cyclophilin, and a domain of a major histocompatibility complex antigen were successfully produced in E. coli as discrete polypeptides by this method.
Ito
W
W
Institute for Comprehensive Medical Science, Fujita Health University, Aichi, Japan.
Kurosawa
Y
Y
D000591:Amino Acid Isomerases / Q000096:biosynthesis / Q000187:drug effects / Q000235:genetics; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002352:Carrier Proteins / Q000096:biosynthesis / Q000187:drug effects / Q000235:genetics; D003001:Cloning, Molecular / Q000379:methods*; D004926:Escherichia coli / Q000235:genetics*; D005796:Genes; D005803:Genes, Immunoglobulin; D005822:Genetic Vectors / Q000235:genetics*; D005982:Glutathione Transferase / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism; D007135:Immunoglobulin Variable Region / Q000096:biosynthesis / Q000187:drug effects / Q000235:genetics; D008285:Major Histocompatibility Complex / Q000235:genetics; D008969:Molecular Sequence Data; D019696:Peptidylprolyl Isomerase; D011994:Recombinant Proteins / Q000096:biosynthesis* / Q000378:metabolism; D012045:Regulatory Sequences, Nucleic Acid / Q000235:genetics; D012549:Schistosoma japonicum / Q000201:enzymology / Q000235:genetics; D022021:Tacrolimus Binding Proteins; D014367:Tryptophan Synthase / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism
D002352:Carrier Proteins; D007135:Immunoglobulin Variable Region; D011994:Recombinant Proteins; D005982:Glutathione Transferase; D014367:Tryptophan Synthase; D000591:Amino Acid Isomerases; D022021:Tacrolimus Binding Proteins; D019696:Peptidylprolyl Isomerase