title: Overlapping transport and chaperone-binding functions within a bacterial twin-arginine signal peptide.
abstract: The twin-arginine translocation (Tat) pathway is a protein targeting system present in many prokaryotes. The physiological role of the Tat pathway is the transmembrane translocation of fully-folded proteins, which are targeted by N-terminal signal peptides bearing conserved SRRxFLK 'twin-arginine' amino acid motifs. In Escherichia coli the majority of Tat targeted proteins bind redox cofactors and it is important that only mature, cofactor-loaded precursors are presented for export. Cellular processes have been unearthed that sequence these events, for example the signal peptide of the periplasmic nitrate reductase (NapA) is bound by a cytoplasmic chaperone (NapD) that is thought to regulate assembly and export of the enzyme. In this work, genetic, biophysical and structural approaches were taken to dissect the interaction between NapD and the NapA signal peptide. A NapD binding epitope was identified towards the N-terminus of the signal peptide, which overlapped significantly with the twin-arginine targeting motif. NMR spectroscopy revealed that the signal peptide adopted a α-helical conformation when bound by NapD, and substitution of single residues within the NapA signal peptide was sufficient to disrupt the interaction. This work provides an increased level of understanding of signal peptide function on the bacterial Tat pathway.
authors.affiliation: College of Life Sciences, University of Dundee, Dundee DD1 5EH, Scotland, UK.
authors.forename: Sabine
authors.initials: S
authors.lastname: Grahl
authors.forename: Julien
authors.initials: J
authors.lastname: Maillard
authors.forename: Chris A E M
authors.initials: CA
authors.lastname: Spronk
authors.forename: Geerten W
authors.initials: GW
authors.lastname: Vuister
authors.forename: Frank
authors.initials: F
authors.lastname: Sargent
mesh_terms: D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D001120:Arginine / Q000378:metabolism*; D001692:Biological Transport; D002352:Carrier Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D004926:Escherichia coli / Q000737:chemistry / Q000201:enzymology / Q000235:genetics / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D008958:Models, Molecular; D018832:Molecular Chaperones / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D008969:Molecular Sequence Data; D050901:Nitrate Reductase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding; D021382:Protein Sorting Signals; D017434:Protein Structure, Tertiary
chemical_list: D002352:Carrier Proteins; D029968:Escherichia coli Proteins; D018832:Molecular Chaperones; C524414:NapD protein, E coli; D021382:Protein Sorting Signals; D001120:Arginine; D050901:Nitrate Reductase; C494404:napA protein, E coli