The Escherichia coli Twin-arginine Translocation Apparatus Incorporates a Distinct Form of TatABC Complex, Spectrum of Modular TatA Complexes and Minor TatAB Complex

Oates, Joanne; Barrett, Claire M.L.; Barnett, James P.; Byrne, Katheryne G.; Bolhuis, Albert; Robinson, Colin

doi:10.1016/j.jmb.2004.11.047

Cited by 104 publications

(143 citation statements)

References 39 publications

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“…This represents a relatively tight peak, suggesting that TatE is rather homogeneous, and calibration of the column shows the complex to be ϳ130 kDa in mass on average, including the detergent micelle. This result is in marked contrast to previous findings on the E. coli TatA complex, which was found to be remarkably heterogeneous and present as complexes ranging in size from ϳ50 kDa to over 500 kDa (10,11). To confirm this point under similar experimental conditions, we expressed E. coli TatA from the plasmid pBAD-A-Strep, where TatA is likewise tagged with a C-terminal Strep-II TM tag, and we purified the protein by Q-Sepharose and Strep-Tactin column using the same protocol (data not shown).…”

Section: Characterization Of Tate Complexes Formed After Overexpressicontrasting

confidence: 99%

“…4. TatA gel filtration elution fractions 18 -25 were subjected to BN gel electrophoresis, and the TatA complexes formed the characteristic ladder of bands observed in an earlier study (10). These bands correspond to separate complexes with estimated sizes ranging from ϳ100 kDa to over 500 kDa, with the gel filtration fractions containing distinct size classes.…”

Section: Characterization Of Tate Complexes Formed After Overexpressimentioning

confidence: 83%

“…Blue Native Polyacrylamide Gel Electrophoresis-Blue native polyacrylamide gel electrophoresis of Tat complexes was performed as described previously (10). Solubilized membranes and purified protein were loaded and separated on a polyacrylamide gradient gel (5-13.5%).…”

Section: Strep-iimentioning

confidence: 99%

“…It is also believed that TatA complexes assemble with the TatABC complex to form a transient "supercomplex," with TatA forming the translocation channel. It has been proposed that the variable size of the TatA complex could be a key feature that enables the system to generate a channel of appropriate diameter for a given substrate (10,11), and electron microscopy studies have shown that TatA complexes do indeed contain potential pores of widely differing diameter (11). Accordingly, TatA is generally considered to form the bulk of the translocation pore.…”

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confidence: 99%

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Structure of TatA Paralog, TatE, Suggests a Structurally Homogeneous Form of Tat Protein Translocase That Transports Folded Proteins of Differing Diameter

Baglieri

Beck

Vasisht

et al. 2012

Journal of Biological Chemistry

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Section: Characterization Of Tate Complexes Formed After Overexpressicontrasting

confidence: 99%

Section: Characterization Of Tate Complexes Formed After Overexpressimentioning

confidence: 83%

Section: Strep-iimentioning

confidence: 99%

mentioning

confidence: 99%

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Structure of TatA Paralog, TatE, Suggests a Structurally Homogeneous Form of Tat Protein Translocase That Transports Folded Proteins of Differing Diameter

Baglieri

Beck

Vasisht

et al. 2012

Journal of Biological Chemistry

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“…1E). Comparable differences in TatA oligomer molecular weights have been reported by others for TatA solubilized in a different detergent (30). Each TatA protomer has a molecular mass of 10.7 kDa, suggesting that successive oligomers differ in composition by more than a single TatA subunit.…”

Section: Resultssupporting

confidence: 57%

The TatA component of the twin-arginine protein transport system forms channel complexes of variable diameter

Gohlke

Pullan

McDevitt

et al. 2005

Proc. Natl. Acad. Sci. U.S.A.

243

274

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The Tat system mediates Sec-independent transport of folded precursor proteins across the bacterial plasma membrane or the chloroplast thylakoid membrane. Tat transport involves distinct high-molecular-weight TatA and TatBC complexes. Here we report the 3D architecture of the TatA complex from Escherichia coli obtained by single-particle electron microscopy and random conical tilt reconstruction. TatA forms ring-shaped structures of variable diameter in which the internal channels are large enough to accommodate known Tat substrate proteins. This morphology strongly supports the proposal that TatA forms the proteinconducting channel of the Tat system. One end of the channel is closed by a lid that might gate access to the channel. On the basis of previous protease accessibility measurements, the lid is likely to be located at the cytoplasmic side of the membrane. The observed variation in TatA diameter suggests a model for Tat transport in which the number of TatA protomers changes to match the size of the channel to the size of the substrate being transported. Such dynamic close packing would provide a mechanism to maintain the membrane permeability barrier during transport.conical tilt reconstruction ͉ electron microscopy ͉ Tat protein transport ͉ three-dimensional structure ͉ twin-arginine signal peptide

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Detection and analysis of protein–protein interactions in organellar and prokaryotic proteomes by native gel electrophoresis: (Membrane) protein complexes and supercomplexes

2006

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It is an essential and challenging task to unravel protein-protein interactions in their actual in vivo context. Native gel systems provide a separation platform allowing the analysis of protein complexes on a rather proteome-wide scale in a single experiment. This review focus on blue-native (BN)-PAGE as the most versatile and successful gel-based approach to separate soluble and membrane protein complexes of intricate protein mixtures derived from all biological sources. BN-PAGE is a charge-shift method with a running pH of 7.5 relying on the gentle binding of anionic CBB dye to all membrane and many soluble protein complexes, leading to separation of protein species essentially according to their size and superior resolution than other fractionation techniques can offer. The closely related colorless-native (CN)-PAGE, whose applicability is restricted to protein species with intrinsic negative net charge, proved to provide an especially mild separation capable of preserving weak protein-protein interactions better than BN-PAGE. The essential conditions determining the success of detecting protein-protein interactions are the sample preparations, e.g. the efficiency/mildness of the detergent solubilization of membrane protein complexes. A broad overview about the achievements of BN- and CN-PAGE studies to elucidate protein-protein interactions in organelles and prokaryotes is presented, e.g. the mitochondrial protein import machinery and oxidative phosphorylation supercomplexes. In many cases, solubilization with digitonin was demonstrated to facilitate an efficient and particularly gentle extraction of membrane protein complexes prone to dissociation by treatment with other detergents. In general, analyses of protein interactomes should be carried out by both BN- and CN-PAGE.

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The Escherichia coli Twin-arginine Translocation Apparatus Incorporates a Distinct Form of TatABC Complex, Spectrum of Modular TatA Complexes and Minor TatAB Complex

Cited by 104 publications

References 39 publications

Structure of TatA Paralog, TatE, Suggests a Structurally Homogeneous Form of Tat Protein Translocase That Transports Folded Proteins of Differing Diameter

Structure of TatA Paralog, TatE, Suggests a Structurally Homogeneous Form of Tat Protein Translocase That Transports Folded Proteins of Differing Diameter

The TatA component of the twin-arginine protein transport system forms channel complexes of variable diameter

Detection and analysis of protein–protein interactions in organellar and prokaryotic proteomes by native gel electrophoresis: (Membrane) protein complexes and supercomplexes

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