Genetic analysis of SecY: additional export-defective mutations and factors affecting their phenotypes.
A number of secY mutants of Escherichia coli showing protein export defects were isolated by a combination of localized mutagenesis and secA-lacZ screening. Most of them were cold sensitive and contained single base substitutions in secY leading to amino acid replacements in various parts of the SecY protein, mainly in the cytoplasmic and the transmembrane domains. A temperature-sensitive mutant with an export defect had the same base substitution as secY24, which was characterized previously. Many cold-sensitive secY mutants exhibited rapid responses to temperature lowering but their apparent defects varied at the permissive temperature. Others exhibited delayed responses to the temperature shift. Some secY mutations, including secY39, interfered with protein export when expressed from a multicopy plasmid, even in the presence of wild-type secY on the chromosome. Such "dominant negative" mutations, including secY-d1, which was studied previously, were all located in either cytoplasmic domain 5 or 6, which is consistent with our previous proposal that the C-terminal region of SecY is important for its function as a protein translocator. We also studied the phenotypes of strains in which one of the secY mutations was combined with the components of the secD operon. Overexpression of secD partially suppressed the secY39 mutation, while overexpression of secF exacerbated the export defects of secY122 and secY125 mutations. Overexpression of "yajC", located within the secD operon, suppressed secY-d1. Although yajC itself proved to be dispensable, its disruption impaired the growth of the secY39 mutant at 42 degrees C. These observations suggest that SecY interacts with SecD, SecF, and the product of yajC.
Taura
T
T
Department of Cell Biology, Kyoto University, Japan.
Akiyama
Y
Y
Ito
K
K
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics*; D001483:Base Sequence; D004926:Escherichia coli / Q000235:genetics; D029968:Escherichia coli Proteins; D008969:Molecular Sequence Data; D009154:Mutation; D000069816:SEC Translocation Channels; D013696:Temperature; D014178:Translocation, Genetic
D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; D000069816:SEC Translocation Channels; C057841:SecY protein, E coli