SecY variants that interfere with Escherichia coli protein export in the presence of normal secY.
As an approach for studying how SecY, an integral membrane protein translocation factor of Escherichia coli, interacts with other protein molecules, we isolated a dominant negative mutation, secY-d1, of the gene carried on a plasmid. The mutant plasmid severely inhibited export of maltose-binding protein and less severely of OmpA, when introduced into sec+ cells. It inhibited growth of secY and secE mutant cells, but not of secA and secD mutant cells or wild-type cells. The mutation deletes three amino acids that should be located at the interface of cytoplasmic domain 5 and transmembrane segment 9. We also found that some SecY-PhoA fusion proteins that lacked carboxy-terminal portions of SecY but retain a region from periplasmic domain 3 to transmembrane segment 7 were inhibitory to protein export. We suggest that these SecY variants are severely defective in catalytic function of SecY, which requires cytoplasmic domain 5 and its carboxy-terminal side, but retain the ability to associate with other molecules of the protein export machinery, which requires the central portion of SecY; they probably exert the 'dominant negative' effects by competing with normal SecY for the formation of active Sec complex. These observations should provide a basis for further genetic analysis of the Sec protein complex in the membrane.
Shimoike
T
T
Department of Cell Biology, Kyoto University, Japan.
Akiyama
Y
Y
Baba
T
T
Taura
T
T
Ito
K
K
D000469:Alkaline Phosphatase / Q000235:genetics; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D001692:Biological Transport / Q000235:genetics; D002455:Cell Division; D004926:Escherichia coli / Q000235:genetics / Q000254:growth & development / Q000378:metabolism*; D029968:Escherichia coli Proteins; D005798:Genes, Bacterial; D005799:Genes, Dominant; D009154:Mutation; D010957:Plasmids; D011993:Recombinant Fusion Proteins / Q000235:genetics; D000069816:SEC Translocation Channels
D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; D011993:Recombinant Fusion Proteins; D000069816:SEC Translocation Channels; C057841:SecY protein, E coli; D000469:Alkaline Phosphatase