title: Towards understanding the Tat translocation mechanism through structural and biophysical studies of the amphipathic region of TatA from Escherichia coli.
abstract: The twin-arginine translocase (Tat) system is used by many bacteria and plants to move folded proteins across the cytoplasmic or thylakoid membrane. In most bacteria, the TatA protein is believed to form a defined pore in the membrane through homo-oligomerization with other TatA protomers. The predicted secondary structure of TatA includes a transmembrane helix, an amphipathic helix, and an unstructured C-terminal region. Here biophysical and structural investigations were performed on a synthetic peptide representing the amphipathic region of TatA (residues 22 to 44, abbreviated TatAH2). The C-terminal region of TatA (residues 44-89) was previously shown to be accessible from both the cytoplasmic and periplasmic sides of the membrane only when the membrane potential was intact, suggesting dependence of its topology on an energized membrane (Chan et al. 2007 Biochemistry 46: 7396-404). Such observation suggests that the TatAH2 region would have unique lipid interactions that may be related to the function of TatA during translocation and thus warranted further investigations. NMR and CD spectroscopy of TatAH2 show that it adopts a predominantly helical structure in a membrane environment while remaining unstructured in aqueous solution. Differential scanning calorimetry studies also reveal that TatAH2 interacts with DPPG lipids but not with DPPC, suggesting that negatively charged phospholipid head groups contribute to the membrane interactions with TatA.
authors.affiliation: Biochemistry Research Cluster, Department of Biological Sciences, University of Calgary, Calgary, Alberta, Canada.
authors.forename: Catherine S
authors.initials: CS
authors.lastname: Chan
authors.forename: Evan F
authors.initials: EF
authors.lastname: Haney
authors.forename: Hans J
authors.initials: HJ
authors.lastname: Vogel
authors.forename: Raymond J
authors.initials: RJ
authors.lastname: Turner
mesh_terms: D015060:1,2-Dipalmitoylphosphatidylcholine / Q000737:chemistry; D001703:Biophysics / Q000379:methods; D002152:Calorimetry, Differential Scanning / Q000379:methods; D002942:Circular Dichroism; D003593:Cytoplasm / Q000378:metabolism; D004926:Escherichia coli / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000737:chemistry* / Q000378:metabolism*; D005452:Fluoresceins / Q000737:chemistry; D008051:Lipid Bilayers / Q000737:chemistry*; D008055:Lipids / Q000737:chemistry; D009682:Magnetic Resonance Spectroscopy / Q000379:methods; D008564:Membrane Potentials; D026901:Membrane Transport Proteins / Q000737:chemistry* / Q000378:metabolism*; D010458:Peptidyl Transferases / Q000737:chemistry*; D010715:Phosphatidylglycerols / Q000737:chemistry; D017510:Protein Folding; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary
chemical_list: D029968:Escherichia coli Proteins; D005452:Fluoresceins; D008051:Lipid Bilayers; D008055:Lipids; D026901:Membrane Transport Proteins; D010715:Phosphatidylglycerols; C499697:TatA protein, E coli; D015060:1,2-Dipalmitoylphosphatidylcholine; D010458:Peptidyl Transferases; C007740:fluorexon; C030345:1,2-dipalmitoylphosphatidylglycerol