title: Identification of the preprotein binding domain of SecA.
abstract: SecA, the preprotein translocase ATPase, has a helicase DEAD motor. To catalyze protein translocation, SecA possesses two additional flexible domains absent from other helicases. Here we demonstrate that one of these "specificity domains" is a preprotein binding domain (PBD). PBD is essential for viability and protein translocation. PBD mutations do not abrogate the basal enzymatic properties of SecA (nucleotide binding and hydrolysis), nor do they prevent SecA binding to the SecYEG protein conducting channel. However, SecA PBD mutants fail to load preproteins onto SecYEG, and their translocation ATPase activity does not become stimulated by preproteins. Bulb and Stem, the two sterically proximal PBD substructures, are physically separable and have distinct roles. Stem binds signal peptides, whereas the Bulb binds mature preprotein regions as short as 25 amino acids. Binding of signal or mature region peptides or full-length preproteins causes distinct conformational changes to PBD and to the DEAD motor. We propose that (a) PBD is a preprotein receptor and a physical bridge connecting bound preproteins to the DEAD motor, and (b) preproteins control the ATPase cycle via PBD.
authors.affiliation: Institute of Molecular Biology and Biotechnology, F.O.R.T.H., University of Crete, P.O. Box 1527, GR-711 10 Iraklio, Crete, Greece.
authors.forename: Efrosyni
authors.initials: E
authors.lastname: Papanikou
authors.forename: Spyridoula
authors.initials: S
authors.lastname: Karamanou
authors.forename: Catherine
authors.initials: C
authors.lastname: Baud
authors.forename: Miriam
authors.initials: M
authors.lastname: Frank
authors.forename: Giorgos
authors.initials: G
authors.lastname: Sianidis
authors.forename: Dimitra
authors.initials: D
authors.lastname: Keramisanou
authors.forename: Charalampos G
authors.initials: CG
authors.lastname: Kalodimos
authors.forename: Andreas
authors.initials: A
authors.lastname: Kuhn
authors.forename: Anastassios
authors.initials: A
authors.lastname: Economou
mesh_terms: D000251:Adenosine Triphosphatases / Q000737:chemistry / Q000502:physiology*; D001412:Bacillus subtilis / Q000378:metabolism; D001426:Bacterial Proteins / Q000502:physiology*; D015374:Biosensing Techniques; D002384:Catalysis; D002462:Cell Membrane / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004926:Escherichia coli / Q000378:metabolism; D029968:Escherichia coli Proteins / Q000737:chemistry / Q000378:metabolism; D006868:Hydrolysis; D007700:Kinetics; D009682:Magnetic Resonance Spectroscopy; D008565:Membrane Proteins / Q000378:metabolism; D026901:Membrane Transport Proteins / Q000502:physiology*; D008957:Models, Genetic; D008958:Models, Molecular; D009154:Mutation; D010455:Peptides / Q000737:chemistry; D040081:Protein Array Analysis; D011485:Protein Binding; D011487:Protein Conformation; D021382:Protein Sorting Signals; D017434:Protein Structure, Tertiary; D021381:Protein Transport; D000069816:SEC Translocation Channels; D000081416:SecA Proteins; D013379:Substrate Specificity; D013696:Temperature
chemical_list: D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; D008565:Membrane Proteins; D026901:Membrane Transport Proteins; D010455:Peptides; D021382:Protein Sorting Signals; D000069816:SEC Translocation Channels; C065493:SecE protein, E coli; C088616:SecG protein, E coli; C057841:SecY protein, E coli; D000251:Adenosine Triphosphatases; D000081416:SecA Proteins