High expression of era gene.
For high production of Era protein, lambda Ea8.5 gene has been chosen from Gene Bank because the Ea8.5 protein contains a sequence of several amino acids at its N-terminus which is identical to that of Era and also because it can be highly expressed in E. coli. The 5'-end sequence of era gene was substituted by a synthetic oligonucleotide which was identical to that of the 5'-end of Ea8.5 gene, so that the transcripts of era gene was provided with a strong translational initiation signal without changing the amino acid sequence of its translational product. Plasmid pCE31, which harbored the recombined era gene under the control of PL promoter, could express very high levels of Era protein, while synthesis of other cellular proteins was nearly shut off during the period of induction. As a result, Era constituted over 80% of the total cellular protein. Electrophoretic pure Era protein with specific guanine nucleotide-binding activity was obtained by a simple procedure including lyzing the cells and washing the pellet of the lysate.
Chen
S
S
Department of Biochemistry, Fourth Military Medical University, Xi'an, Shaanxi, China.
Court
D L
DL
D001426:Bacterial Proteins / Q000096:biosynthesis / Q000235:genetics* / Q000302:isolation & purification; D001483:Base Sequence; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism / Q000648:ultrastructure; D029968:Escherichia coli Proteins; D019204:GTP-Binding Proteins / Q000096:biosynthesis / Q000235:genetics* / Q000302:isolation & purification; D005798:Genes, Bacterial; D008854:Microscopy, Electron; D008969:Molecular Sequence Data; D010957:Plasmids; D016601:RNA-Binding Proteins; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics / Q000302:isolation & purification
D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; D016601:RNA-Binding Proteins; D011994:Recombinant Proteins; C068027:era protein, E coli; D019204:GTP-Binding Proteins