title: Functional and structural characterization of the minimal Sec translocase of the hyperthermophile Thermotoga maritima.
abstract: The genome of the hyperthermophilic bacterium Thermotoga maritima contains the genes that encode core subunits of the protein translocase, a complex consisting of the molecular motor SecA and the protein conducting pore SecYE. In addition, we identified an erroneous sequence in the genome encoding for a putative secG gene. The genes of the T. maritima translocase subunits were overexpressed in Escherichia coli and purified to homogeneity. T. maritima SecA showed a basal thermostable ATPase activity that was stimulated up to 4-fold by phospholipids with an optimum at 74 degrees C. Membrane vesicles and proteoliposomes containing SecYE or SecYEG supported 2- to 4-fold stimulation of the precursor dependent SecA ATPase activity. Imaging of small two-dimensional crystals of the SecYE complex using electron microscopy showed square-shaped particles with a side-length of about 6 nm. These results demonstrate that in T. maritima a highly thermostable translocase complex is operational.
authors.affiliation: Department of Molecular Microbiology, Groningen Biomolecular Sciences and Biotechnology Institute, University of Groningen, Haren, The Netherlands.
authors.forename: Monika G
authors.initials: MG
authors.lastname: Pretz
authors.forename: Hervé
authors.initials: H
authors.lastname: Remigy
authors.forename: Jelto
authors.initials: J
authors.lastname: Swaving
authors.forename: Sonja V
authors.initials: SV
authors.lastname: Albers
authors.forename: Victoria G
authors.initials: VG
authors.lastname: Garrido
authors.forename: Mohamed
authors.initials: M
authors.lastname: Chami
authors.forename: Andreas
authors.initials: A
authors.lastname: Engel
authors.forename: Arnold J M
authors.initials: AJ
authors.lastname: Driessen
mesh_terms: D000251:Adenosine Triphosphatases / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D003001:Cloning, Molecular; D003460:Crystallization; D026901:Membrane Transport Proteins / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D008854:Microscopy, Electron; D008969:Molecular Sequence Data; D046912:Multiprotein Complexes / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D021122:Protein Subunits / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D021381:Protein Transport; D000069816:SEC Translocation Channels; D000081416:SecA Proteins; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D013696:Temperature; D020124:Thermotoga maritima / Q000201:enzymology* / Q000235:genetics
chemical_list: D001426:Bacterial Proteins; D026901:Membrane Transport Proteins; D046912:Multiprotein Complexes; D021122:Protein Subunits; D000069816:SEC Translocation Channels; D000251:Adenosine Triphosphatases; D000081416:SecA Proteins