title: FtsH, a membrane-bound ATPase, forms a complex in the cytoplasmic membrane of Escherichia coli.
abstract: The FtsH (HflB) protein of Escherichia coli is integrated into the membrane with two N-terminally located transmembrane segments, while its large cytoplasmic domain is homologous to the AAA family of ATPases. The previous studies on dominant negative ftsH mutants raised a possibility that FtsH functions in multimeric states. We found that FtsH was eluted at fractions corresponding to a larger molecular weight than expected from monomeric structure in size-exclusion chromatography. Moreover, treatment of membranes or their detergent extracts with a cross-linker, dithiobis(succinimidyl propionate), yielded cross-linked products of FtsH. To dissect possible FtsH complex, we constructed an FtsH derivative with c-Myc epitope at its C terminus (FtsH-His6-Myc). When membranes prepared from cells in which FtsH-His6-Myc was overproduced together with the normal FtsH were treated with the cross-linker, intact FtsH and in vitro degradation products of FtsH-His6-Myc without the tag were cross-linked with the tagged FtsH protein. Co-immunoprecipitation experiments confirmed the interaction between the FtsH molecules. To identify regions of FtsH required or sufficient for this interaction, we constructed chimeric proteins between FtsH and EnvZ, a protein with a similar topological arrangement, by exchanging their corresponding domains. We found that only the FtsH-EnvZ hybrid protein with an FtsH-derived membrane anchoring domain and an EnvZ-derived cytoplasmic domain caused a dominant ftsH phenotype and was cross-linked with FtsH. We suggest that the N-terminal transmembrane region of FtsH mediates directly the interaction between the FtsH subunits.
authors.affiliation: Department of Cell Biology, Kyoto University, Japan.
authors.forename: Y
authors.initials: Y
authors.lastname: Akiyama
authors.forename: T
authors.initials: T
authors.lastname: Yoshihisa
authors.forename: K
authors.initials: K
authors.lastname: Ito
mesh_terms: D049069:ATP-Dependent Proteases; D000251:Adenosine Triphosphatases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D000595:Amino Acid Sequence; D001425:Bacterial Outer Membrane Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D001483:Base Sequence; D001665:Binding Sites; D002462:Cell Membrane / Q000378:metabolism; D003432:Cross-Linking Reagents; D003593:Cytoplasm / Q000378:metabolism; D017931:DNA Primers / Q000235:genetics; D004269:DNA, Bacterial / Q000235:genetics; D000939:Epitopes / Q000737:chemistry; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D029968:Escherichia coli Proteins; D008565:Membrane Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D015394:Molecular Structure; D009097:Multienzyme Complexes; D010641:Phenotype; D011233:Precipitin Tests; D011487:Protein Conformation; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism
chemical_list: D001425:Bacterial Outer Membrane Proteins; D001426:Bacterial Proteins; D003432:Cross-Linking Reagents; D017931:DNA Primers; D004269:DNA, Bacterial; D000939:Epitopes; D029968:Escherichia coli Proteins; D008565:Membrane Proteins; D009097:Multienzyme Complexes; D011993:Recombinant Fusion Proteins; C057843:envZ protein, E coli; D049069:ATP-Dependent Proteases; C513383:FtsH protein, E coli; D000251:Adenosine Triphosphatases