Expression of chimeric ras protein with OmpF signal peptide in Escherichia coli: localization of OmpF fusion protein in the inner membrane.
The ras gene was fused with the DNA sequence of OmpF signal peptide or with the DNA sequence of OmpF signal peptide plus the amino terminal portion of the OmpF gene. They were placed in plasmids together with the bacteriophage lambda PL promoter. These plasmids were introduced into Escherichia coli strain K-12 and the OmpF signal peptide fusion proteins were expressed. These fusion proteins were identified as 29.0 and 30.0 kDa proteins. However, processed products of these proteins were not found in the extract. The fusion proteins were localized mostly in the cytoplasm and the inner membrane, but none of them was secreted into the periplasmic space. On the other hand, the ras protein alone was found in the cytoplasm and not in the inner membrane. Viable counts of E. coli harbouring these plasmids decreased when these fused proteins were induced. Induction of the ras protein alone did not harm cells. These observations suggest that insertion of the heterologous proteins into the inner membrane may cause the bactericidal effect.
Yamamoto
T
T
Research Institute for Molecular Genetics, Tsumura & Co., Ibaraki, Japan.
Okawa
N
N
Endo
T
T
Kaji
A
A
D000595:Amino Acid Sequence; D001425:Bacterial Outer Membrane Proteins / Q000032:analysis / Q000235:genetics*; D010582:Bacteriophage lambda / Q000235:genetics; D001483:Base Sequence; D002462:Cell Membrane / Q000201:enzymology / Q000502:physiology / Q000648:ultrastructure; D002678:Chimera; D004926:Escherichia coli / Q000235:genetics* / Q000254:growth & development; D011905:Genes, ras; D002479:Inclusion Bodies / Q000201:enzymology; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010957:Plasmids; D021382:Protein Sorting Signals / Q000032:analysis / Q000235:genetics*; D016283:Proto-Oncogene Proteins p21(ras) / Q000032:analysis / Q000235:genetics*; D011993:Recombinant Fusion Proteins / Q000032:analysis; D013385:Succinate Dehydrogenase / Q000378:metabolism
D001425:Bacterial Outer Membrane Proteins; D021382:Protein Sorting Signals; D011993:Recombinant Fusion Proteins; D013385:Succinate Dehydrogenase; D016283:Proto-Oncogene Proteins p21(ras)