title: FtsH (HflB) is an ATP-dependent protease selectively acting on SecY and some other membrane proteins.
abstract: The FtsH protein is a membrane-bound ATPase of Escherichia coli that was proposed to be involved in membrane protein assembly as well as degradation of some unstable proteins. SecY, a subunit of protein translocase, is FtsH dependently degraded in vivo when it fails to associate with its partner (the SecE protein). We constructed a series of mutants in which mutations were introduced into conserved residues in the two ATP binding consensus sequences or the zinc binding sequence of FtsH. We purified wild-type and mutant FtsH proteins by making use of a polyhistidine tag attached to their carboxyl termini. Complementation analysis and ATPase activity assays in vitro indicated that, of the two sets of ATP binding sequence motifs, the one located C-terminally (A1) is essential for ATPase activity and in vivo functioning of FtsH. Wild-type FtsH protein degraded purified SecY in an ATP hydrolysis-dependent manner in vitro. Mutant proteins without ATPase activity were inactive in proteolysis. A zinc binding motif mutant showed a decreased proteolytic activity. SecY and FtsH were cross-linkable with each other in the membrane, provided that FtsH had an ATPase-inactivating mutation. These results demonstrate that FtsH binds to and degrades SecY, its A1 motif and the zinc binding motif being important for the proteolytic activity. FtsH-dependent proteolysis was also demonstrated for SecY in crude membrane extracts, whereas a majority of other membrane proteins were not degraded, indicating that FtsH has high selectivity in protein degradation.
authors.affiliation: Department of Cell Biology, Institute for Virus Research, Kyoto University, Kyoto 606-01, Japan. yakiyama@virus.kyoto-u.ac.jp
authors.forename: Y
authors.initials: Y
authors.lastname: Akiyama
authors.forename: A
authors.initials: A
authors.lastname: Kihara
authors.forename: H
authors.initials: H
authors.lastname: Tokuda
authors.forename: K
authors.initials: K
authors.lastname: Ito
mesh_terms: D049069:ATP-Dependent Proteases; D000251:Adenosine Triphosphatases / Q000378:metabolism*; D000255:Adenosine Triphosphate / Q000378:metabolism*; D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000378:metabolism*; D001665:Binding Sites; D004591:Electrophoresis, Polyacrylamide Gel; D010450:Endopeptidases / Q000378:metabolism*; D029968:Escherichia coli Proteins; D008565:Membrane Proteins / Q000378:metabolism*; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010641:Phenotype; D010957:Plasmids / Q000378:metabolism; D000069816:SEC Translocation Channels
chemical_list: D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; D008565:Membrane Proteins; D000069816:SEC Translocation Channels; C057841:SecY protein, E coli; D000255:Adenosine Triphosphate; D010450:Endopeptidases; D049069:ATP-Dependent Proteases; C513383:FtsH protein, E coli; D000251:Adenosine Triphosphatases