scholarly journals Defining the DmsA Signal Sequence Interaction with DmsD And TatBC

2019 ◽  
Author(s):  
Deepanjan Ghosh ◽  
Sureshkumar Ramasamy
Keyword(s):  
Complex Formation ◽  
Signal Sequence ◽  
Factor Loading ◽  
Binding Studies ◽  
Redox Enzyme ◽  
Twin Arginine Translocation ◽  
Membrane Complex ◽  
Monomeric Complex ◽  
Tat Pathway ◽  
Complex Forms

AbstractRedox – enzyme maturation proteins (REMP) ensure co-factor loading and folding of proteins targeted to the twin-arginine translocation (Tat) pathway. The details of the interaction of a REMP with the corresponding signal sequence of its substrate are not well understood. Here, we demonstrate the features of the signal sequence for the Tat substrate DmsA (ssDmsA) responsible for complex formation with its REMP, DmsD, and with the Tat membrane complex TatB & TatC (TatBC). A heterologously expressed ssDmsA/DmsD complex forms two stochiometric populations corresponding to monomeric and dimeric forms of the complex. The monomeric complex has a higher affinity for the TatBC complex than the dimeric, which imply higher level regulation process to ensure the maturation of protein before translocation. Results from various binding studies yielded the shortest signal peptide required for ssDmsA/DmsA interaction and the region responsible for the TatBC interaction. Further experiments like alanine scanning in this peptide highlight the possible residues that are essential for this complex formation.

scholarly journals Role of the Twin-Arginine Translocation Pathway in Staphylococcus

2009 ◽  
Vol 191 (19) ◽  
pp. 5921-5929 ◽  
Author(s):  
Lalitha Biswas ◽  
Raja Biswas ◽  
Christiane Nerz ◽  
Knut Ohlsen ◽  
Martin Schlag ◽  
...  
Keyword(s):  
Functional Unit ◽  
Signal Sequence ◽  
Protein A ◽  
Mouse Kidney ◽  
Deletion Mutants ◽  
Twin Arginine Translocation ◽  
High Affinity ◽  
Tat Pathway ◽  
First Time

ABSTRACT In Staphylococcus, the twin-arginine translocation (Tat) pathway is present only in some species and is composed of TatA and TatC. The tatAC operon is associated with the fepABC operon, which encodes homologs to an iron-binding lipoprotein, an iron-dependent peroxidase (FepB), and a high-affinity iron permease. The FepB protein has a typical twin-arginine (RR) signal peptide. The tat and fep operons constitute an entity that is not present in all staphylococcal species. Our analysis was focused on Staphylococcus aureus and S. carnosus strains. Tat deletion mutants (ΔtatAC) were unable to export active FepB, indicating that this enzyme is a Tat substrate. When the RR signal sequence from FepB was fused to prolipase and protein A, their export became Tat dependent. Since no other protein with a Tat signal could be detected, the fepABC-tatAC genes comprise not only a genetic but also a functional unit. We demonstrated that FepABC drives iron import, and in a mouse kidney abscess model, the bacterial loads of ΔtatAC and Δtat-fep mutants were decreased. For the first time, we show that the Tat pathway in S. aureus is functional and serves to translocate the iron-dependent peroxidase FepB.

Identification of Streptomyces lividans proteins secreted by the twin-arginine translocation pathway following growth with different carbon sources

2008 ◽  
Vol 54 (7) ◽  
pp. 549-558 ◽  
Author(s):  
Julien Guimond ◽  
Rolf Morosoli
Keyword(s):  
Signal Sequence ◽  
Streptomyces Coelicolor ◽  
Carbon Sources ◽  
Streptomyces Lividans ◽  
Extracellular Proteins ◽  
Soil Extracts ◽  
Twin Arginine Translocation ◽  
Tat Pathway ◽  
Close Proximity ◽  
Dependent Protein

Genome-based signal peptide predictions classified Streptomyces coelicolor as the microorganism that secretes the most proteins through the twin-arginine translocation (Tat)-dependent secretion pathway. Availability of a ΔtatC mutant of the closely related strain Streptomyces lividans impaired Tat-dependent protein secretion and enabled identification of many extracellular proteins that are secreted via the Tat pathway. Proteomic techniques were applied to analyze proteins from the supernatants of log-phase cultures. Since the bacterial secretome depends mainly on the carbon sources available during growth, xylose, glucose, chitin, and soil extracts were used. A total of 63 proteins were identified, among which 7 were predicted by the TATscan program, and 20 were not predicted but contained a potential Tat signal motif. Thirteen proteins having no signal sequence could be co-transported by Tat-dependent proteins because the genes that encode these proteins are in close proximity in the genome. Finally, the presence of 23 proteins lacking signal peptides was difficult to explain. More secreted proteins could be identified as Tat substrates in varying carbon sources.

scholarly journals Phage Shock Protein PspA of Escherichia coli Relieves Saturation of Protein Export via the Tat Pathway

2004 ◽  
Vol 186 (2) ◽  
pp. 366-373 ◽  
Author(s):  
Matthew P. DeLisa ◽  
Philip Lee ◽  
Tracy Palmer ◽  
George Georgiou
Keyword(s):  
Genomic Library ◽  
Protein Translocation ◽  
Signal Sequence ◽  
Protein A ◽  
Homologous Proteins ◽  
Gene Encoding ◽  
Protein Precursor ◽  
Twin Arginine Translocation ◽  
Tat Pathway

ABSTRACT Overexpression of either heterologous or homologous proteins that are routed to the periplasm via the twin-arginine translocation (Tat) pathway results in a block of export and concomitant accumulation of the respective protein precursor in the cytoplasm. Screening of a plasmid-encoded genomic library for mutants that confer enhanced export of a TorA signal sequence (ssTorA)-GFP-SsrA fusion protein, and thus result in higher cell fluorescence, yielded the pspA gene encoding phage shock protein A. Coexpression of pspA relieved the secretion block observed with ssTorA-GFP-SsrA or upon overexpression of the native Tat proteins SufI and CueO. A similar effect was observed with the Synechocystis sp. strain PCC6803 PspA homologue, VIPP1, indicating that the role of PspA in Tat export may be phylogenetically conserved. Mutations in Tat components that completely abolish export result in a marked induction of PspA protein synthesis, consistent with its proposed role in enhancing protein translocation via Tat.

scholarly journals Genetic Analysis of Pathway Specificity during Posttranslational Protein Translocation across the Escherichia coli Plasma Membrane

2003 ◽  
Vol 185 (9) ◽  
pp. 2811-2819 ◽  
Author(s):  
Natascha Blaudeck ◽  
Peter Kreutzenbeck ◽  
Roland Freudl ◽  
Georg A. Sprenger
Keyword(s):  
Escherichia Coli ◽  
Signal Peptide ◽  
Protein Translocation ◽  
Alternative Possibilities ◽  
Amino Acid Residues ◽  
Sec Pathway ◽  
Twin Arginine Translocation ◽  
Tat Pathway ◽  
Arginine Residues ◽  
Dependent Protein

ABSTRACT In Escherichia coli, the SecB/SecA branch of the Sec pathway and the twin-arginine translocation (Tat) pathway represent two alternative possibilities for posttranslational translocation of proteins across the cytoplasmic membrane. Maintenance of pathway specificity was analyzed using a model precursor consisting of the mature part of the SecB-dependent maltose-binding protein (MalE) fused to the signal peptide of the Tat-dependent TorA protein. The TorA signal peptide selectively and specifically directed MalE into the Tat pathway. The characterization of a spontaneous TorA signal peptide mutant (TorA*), in which the two arginine residues in the c-region had been replaced by one leucine residue, showed that the TorA*-MalE mutant precursor had acquired the ability for efficiently using the SecB/SecA pathway. Despite the lack of the “Sec avoidance signal,” the mutant precursor was still capable of using the Tat pathway, provided that the kinetically favored Sec pathway was blocked. These results show that the h-region of the TorA signal peptide is, in principle, sufficiently hydrophobic for Sec-dependent protein translocation, and therefore, the positively charged amino acid residues in the c-region represent a major determinant for Tat pathway specificity. Tat-dependent export of TorA-MalE was significantly slower in the presence of SecB than in its absence, showing that SecB can bind to this precursor despite the presence of the Sec avoidance signal in the c-region of the TorA signal peptide, strongly suggesting that the function of the Sec avoidance signal is not the prevention of SecB binding; rather, it must be exerted at a later step in the Sec pathway.

scholarly journals Erratum to: Novel GFP Expression Using a Short N-Terminal Polypeptide through the Defined Twin-Arginine Translocation (Tat) Pathway

2011 ◽  
Vol 32 (5) ◽  
pp. 489-489
Author(s):  
Sang Jun Lee ◽  
Yun Hee Han ◽  
Young Ok Kim ◽  
Bo Hye Nam ◽  
Hee Jeong Kong
Keyword(s):  
Gfp Expression ◽  
Twin Arginine Translocation ◽  
Tat Pathway

scholarly journals Dual targeting of TatA points to a chloroplast-like Tat pathway in plant mitochondria

2020 ◽  
Author(s):  
Bationa Bennewitz ◽  
Mayank Sharma ◽  
Franzisca Tannert ◽  
Ralf Bernd Klösgen
Keyword(s):  
Plant Mitochondria ◽  
Common Mechanism ◽  
Twin Arginine Translocation ◽  
Dual Targeting ◽  
In Organello ◽  
Highly Sensitive ◽  
Tat Pathway ◽  
Electron Transport Chains ◽  
Membrane Bound ◽  
Folded Proteins

AbstractThe biogenesis of membrane-bound electron transport chains requires membrane translocation pathways for folded proteins carrying complex cofactors, like the Rieske Fe/S proteins. Two independent systems were developed during evolution, namely the Twin-arginine translocation (Tat) pathway, which is present in bacteria and chloroplasts, and the Bcs1 pathway found in mitochondria of yeast and mammals. Mitochondria of plants carry a Tat-like pathway which was hypothesized to operate with only two subunits, a TatB-like protein and a TatC homolog (OrfX), but lacking TatA. Here we show that the nuclearly encoded TatA has dual targeting properties, i.e., it can be imported into both, chloroplasts and mitochondria. Dual targeting of TatA was observed with in organello experiments employing isolated chloroplasts and mitochondria as well as after transient expression of suitable reporter constructs in epidermal leaf cells. The extent of transport of these constructs into mitochondria of transiently transformed leaf cells was relatively low, causing a demand for highly sensitive methods to be detected, like the sasplitGFP approach. Yet, the dual import of TatA into mitochondria and chloroplasts observed here points to a common mechanism of Tat transport for folded proteins within both endosymbiotic organelles.

scholarly journals The Twin-Arginine Translocation System Is Important for Stress Resistance and Virulence of Brucella melitensis

2020 ◽  
Vol 88 (11) ◽  
Author(s):  
Xin Yan ◽  
Sen Hu ◽  
Yan Yang ◽  
Da Xu ◽  
Huoming Li ◽  
...  
Keyword(s):  
Ex Vivo ◽  
Brucella Melitensis ◽  
Cell Envelope ◽  
Critical Role ◽  
Sodium Dodecyl ◽  
Methionine Sulfoxide Reductase ◽  
Content Type ◽  
Twin Arginine Translocation ◽  
Tat Pathway

ABSTRACT Brucella, the causative agent of brucellosis, is a stealthy intracellular pathogen that is highly pathogenic to a range of mammals, including humans. The twin-arginine translocation (Tat) pathway transports folded proteins across the cytoplasmic membrane and has been implicated in virulence in many bacterial pathogens. However, the roles of the Tat system and related substrates in Brucella remain unclear. We report here that disruption of Tat increases the sensitivity of Brucella melitensis M28 to the membrane stressor sodium dodecyl sulfate (SDS), indicating cell envelope defects, as well as to EDTA. In addition, mutating Tat renders M28 bacteria more sensitive to oxidative stress caused by H2O2. Further, loss of Tat significantly attenuates B. melitensis infection in murine macrophages ex vivo. Using a mouse model for persistent infection, we demonstrate that Tat is required for full virulence of B. melitensis M28. Genome-wide in silico prediction combined with an in vivo amidase reporter assay indicates that at least 23 proteins are authentic Tat substrates, and they are functionally categorized into solute-binding proteins, oxidoreductases, cell envelope biosynthesis enzymes, and others. A comprehensive deletion study revealed that 6 substrates contribute significantly to Brucella virulence, including an l,d-transpeptidase, an ABC transporter solute-binding protein, and a methionine sulfoxide reductase. Collectively, our work establishes that the Tat pathway plays a critical role in Brucella virulence.

Circulating IgG-LD complex, dissociable by addition of NAD+.

1982 ◽  
Vol 28 (1) ◽  
pp. 236-239 ◽  
Author(s):  
F Gorus ◽  
W Aelbrecht ◽  
B Van Camp
Keyword(s):  
Lactate Dehydrogenase ◽  
Autoimmune Disease ◽  
Complex Formation ◽  
Affinity Chromatography ◽  
Conformational Changes ◽  
Active Site ◽  
Nicotinamide Adenine Dinucleotide ◽  
Binding Sites ◽  
Gel Filtration ◽  
Binding Studies

Abstract Macromolecular LD (lactate dehydrogenase, EC 1.1.1.27) was present in the serum of a patient suffering from idiopathic fibrosis of the lung and presenting signs of autoimmune disease. By using gel filtration and affinity chromatography techniques, the vast majority of the patient's serum LD activity was shown to consist of LD-IgG complexes, which dissociated in the presence of added nicotinamide adenine dinucleotide (NAD+). Binding studies with tritiated NAD+ indicated that complex formation was not ascribable to a lack of circulating cofactor. The most likely explanation for the complex formation was the existence of LD binding sites on IgG molecules. The disruption of the complex by NAD+ might be explained by a competition between IgG molecules and NAD+ for the LD active site or by conformational changes induced in the LD molecules on binding of NAD+.

scholarly journals The Glove-like Structure of the Conserved Membrane Protein TatC Provides Insight into Signal Sequence Recognition in Twin-Arginine Translocation

Structure ◽  
2013 ◽  
Vol 21 (5) ◽  
pp. 777-788 ◽  
Author(s):  
Sureshkumar Ramasamy ◽  
Ravinder Abrol ◽  
Christian J.M. Suloway ◽  
William M. Clemons
Keyword(s):  
Membrane Protein ◽  
Signal Sequence ◽  
Twin Arginine Translocation ◽  
Sequence Recognition ◽  
Insight Into
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