title: Essential cytoplasmic domains in the Escherichia coli TatC protein.
abstract: The twin-arginine translocation (Tat) system mediates the transport of proteins across the bacterial plasma membrane and chloroplast thylakoid membrane. Operating in parallel with Sec-type systems in these membranes, the Tat system is completely different in both structural and mechanistic terms, and is uniquely able to catalyze the translocation of fully folded proteins across coupled membranes. TatC is an essential, multispanning component that has been proposed to form part of the binding site for substrate precursor proteins. In this study we have tested the importance of conserved residues on the periplasmic and cytoplasmic face of the Escherichia coli protein. We find that many of the mutations on the cytoplasmic face have little or no effect. However, substitution at several positions in the extreme N-terminal cytoplasmic region or the predicted first cytoplasmic loop lead to a significant or complete loss of Tat-dependent export. The mutated strains are unable to grow anaerobically on trimethylamine N-oxide minimal media and are unable to export trimethylamine-N-oxide reductase (TorA). The same mutants are completely unable to export a chimeric protein, comprising the TorA signal peptide linked to green fluorescent protein, indicating that translocation is blocked rather than cofactor insertion into the TorA mature protein. The data point to two essential cytoplasmic domains on the TatC protein that are essential for export.
authors.affiliation: Department of Biological Sciences, University of Warwick, Coventry CV4 7AL, United Kingdom.
authors.forename: Stuart C H
authors.initials: SC
authors.lastname: Allen
authors.forename: Claire M L
authors.initials: CM
authors.lastname: Barrett
authors.forename: Nicola
authors.initials: N
authors.lastname: Ray
authors.forename: Colin
authors.initials: C
authors.lastname: Robinson
mesh_terms: D000595:Amino Acid Sequence; D017360:Arabidopsis / Q000378:metabolism; D001120:Arginine / Q000737:chemistry; D001665:Binding Sites; D002462:Cell Membrane / Q000378:metabolism; D003593:Cytoplasm / Q000737:chemistry*; D004926:Escherichia coli / Q000378:metabolism*; D049452:Green Fluorescent Proteins; D008164:Luminescent Proteins / Q000378:metabolism; D008565:Membrane Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D009247:NADH, NADPH Oxidoreductases / Q000378:metabolism; D000587:Oxidoreductases Acting on CH-NH Group Donors; D010089:Oxidoreductases, N-Demethylating / Q000737:chemistry / Q000378:metabolism; D010940:Plant Proteins; D010957:Plasmids / Q000378:metabolism; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D021381:Protein Transport; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D013347:Subcellular Fractions
chemical_list: D008164:Luminescent Proteins; D008565:Membrane Proteins; D010940:Plant Proteins; D011993:Recombinant Fusion Proteins; C432107:TatC protein, plant; D049452:Green Fluorescent Proteins; D001120:Arginine; C021567:methylamine dehydrogenase; D000587:Oxidoreductases Acting on CH-NH Group Donors; D010089:Oxidoreductases, N-Demethylating; C023186:trimethylamine dehydrogenase; D009247:NADH, NADPH Oxidoreductases