Cytosolic factor purified from Escherichia coli is necessary and sufficient for the export of a preprotein and is a homotetramer of SecB.
We have purified to homogeneity a cytosolic factor from Escherichia coli that is required for the translocation of a preprotein into inverted vesicles of the E. coli plasma membrane. The preprotein used is a precursor of mutant maltose-binding protein. This mutant contains alterations of the carboxyl terminus. Unlike the precursor for wild-type maltose-binding protein, the mutant precursor does not acquire a protease-resistant conformation after synthesis and retains posttranslational translocation competence. The purified cytosolic factor, added posttranslationally, is necessary and sufficient to yield virtually 100% translocation of the mutant precursor into inverted vesicles. The purified factor amounts to 0.08% of the cytosolic proteins and is a 64-kDa tetramer consisting of four identical 16-kDa subunits. Amino-terminal sequence analysis revealed that it is identical to the secB gene product. The purified SecB homotetramer is part of a larger 150-kDa complex that represents the "export" factor activity. During purification, the export factor activity dissociates into a 64-kDa SecB homotetramer and unidentified component(s). For the posttranslational integration of another preprotein, the precursor for the lamB gene product, into inverted vesicles, the 64-kDa SecB homotetramer is also required but additional factor(s) makes integration more efficient.
Watanabe
M
M
Laboratory of Cell Biology, Howard Hughes Medical Institute, Rockefeller University, New York, NY 10021.
Blobel
G
G
D018528:ATP-Binding Cassette Transporters; D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000302:isolation & purification / Q000502:physiology*; D002352:Carrier Proteins / Q000378:metabolism*; D002845:Chromatography; D003600:Cytosol / Q000032:analysis; D004926:Escherichia coli / Q000032:analysis / Q000502:physiology*; D029968:Escherichia coli Proteins; D005798:Genes, Bacterial; D066298:In Vitro Techniques; D058619:Maltose-Binding Proteins; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009004:Monosaccharide Transport Proteins; D011499:Protein Processing, Post-Translational
D018528:ATP-Binding Cassette Transporters; D001426:Bacterial Proteins; D002352:Carrier Proteins; D029968:Escherichia coli Proteins; D058619:Maltose-Binding Proteins; D009004:Monosaccharide Transport Proteins; C461599:maltose transport system, E coli