%0 Journal Article %1 frobelSurfaceexposedDomainsTatB2019 %A Fröbel, Julia %A Blümmel, Anne-Sophie %A Drepper, Friedel %A Warscheid, Bettina %A Müller, Matthias %C United States %D 2019 %J The Journal of biological chemistry %K (MS),membrane Acid Amino Binding/physiology,protein Folding,Protein Liquid/methods,Escherichia Mass Membrane/metabolism,Chromatography Molecular,Protein Proteins/*metabolism/physiology,Escherichia Proteins/*metabolism/physiology,Models Proteins/metabolism,Cell Proteins/metabolism,membrane Relationship,structure-function,Tandem Sequence/genetics,Binding Signals/genetics,protein Sites/genetics,Carrier Sorting Spectrometry/methods,Tat System/*metabolism/physiology,zero Transport Transport/physiology,Structure-Activity assembly,TatB,TatBC coli coli/metabolism,LC-MS/MS,mass complex,protein complex,to_read,translocation,twin-arginine cross-linker cross-linking,protein export,Protein protein,Membrane space spectrometry trafficking,membrane translocase translocation,Protein translocation,Twin-Arginine-Translocation transport,Membrane %N 38 %P 13902--13914 %R 10.1074/jbc.RA119.009298 %T Surface-Exposed Domains of TatB Involved in the Structural and Functional Assembly of the Tat Translocase in Escherichia Coli. %V 294 %X Twin-arginine-dependent translocases transport folded proteins across bacterial, archaeal, and chloroplast membranes. Upon substrate binding, they assemble from hexahelical TatC and single-spanning TatA and TatB membrane proteins. Although structural and functional details of individual Tat subunits have been reported previously, the sequence and dynamics of Tat translocase assembly remain to be determined. Employing the zero-space cross-linker N,N'-dicyclohexylcarbodiimide (DCCD) in combination with LC-MS/MS, we identified as yet unknown intra- and intermolecular contact sites of TatB and TatC. In addition to their established intramembrane binding sites, both proteins were thus found to contact each other through the soluble N terminus of TatC and the interhelical linker region around the conserved glutamyl residue Glu(49) of TatB from Escherichia coli Functional analyses suggested that by interacting with the TatC N terminus, TatB improves the formation of a proficient substrate recognition site of TatC. The Glu(49) region of TatB was found also to contact distinct downstream sites of a neighboring TatB molecule and to thereby mediate oligomerization of TatB within the TatBC receptor complex. Finally, we show that global DCCD-mediated cross-linking of TatB and TatC in membrane vesicles or, alternatively, creating covalently linked TatC oligomers prevents TatA from occupying a position close to the TatBC-bound substrate. Collectively, our results are consistent with a circular arrangement of the TatB and TatC units within the TatBC receptor complex and with TatA entering the interior TatBC-binding cavity through lateral gates between TatBC protomers.

@article{frobelSurfaceexposedDomainsTatB2019, abstract = {Twin-arginine-dependent translocases transport folded proteins across bacterial, archaeal, and chloroplast membranes. Upon substrate binding, they assemble from hexahelical TatC and single-spanning TatA and TatB membrane proteins. Although structural and functional details of individual Tat subunits have been reported previously, the sequence and dynamics of Tat translocase assembly remain to be determined. Employing the zero-space cross-linker N,N'-dicyclohexylcarbodiimide (DCCD) in combination with LC-MS/MS, we identified as yet unknown intra- and intermolecular contact sites of TatB and TatC. In addition to their established intramembrane binding sites, both proteins were thus found to contact each other through the soluble N terminus of TatC and the interhelical linker region around the conserved glutamyl residue Glu(49) of TatB from Escherichia coli Functional analyses suggested that by interacting with the TatC N terminus, TatB improves the formation of a proficient substrate recognition site of TatC. The Glu(49) region of TatB was found also to contact distinct downstream sites of a neighboring TatB molecule and to thereby mediate oligomerization of TatB within the TatBC receptor complex. Finally, we show that global DCCD-mediated cross-linking of TatB and TatC in membrane vesicles or, alternatively, creating covalently linked TatC oligomers prevents TatA from occupying a position close to the TatBC-bound substrate. Collectively, our results are consistent with a circular arrangement of the TatB and TatC units within the TatBC receptor complex and with TatA entering the interior TatBC-binding cavity through lateral gates between TatBC protomers.}, added-at = {2024-05-17T13:01:35.000+0200}, address = {United States}, author = {Fr{\"o}bel, Julia and Bl{\"u}mmel, Anne-Sophie and Drepper, Friedel and Warscheid, Bettina and M{\"u}ller, Matthias}, biburl = {https://www.bibsonomy.org/bibtex/27f151ade56e647c345315f632ae1fe65/warscheidlab}, copyright = {{\copyright} 2019 Fr{\"o}bel et al.}, doi = {10.1074/jbc.RA119.009298}, interhash = {c4bcf2bf5e93f5012e4dbdabf03322d5}, intrahash = {7f151ade56e647c345315f632ae1fe65}, issn = {1083-351X 0021-9258}, journal = {The Journal of biological chemistry}, keywords = {(MS),membrane Acid Amino Binding/physiology,protein Folding,Protein Liquid/methods,Escherichia Mass Membrane/metabolism,Chromatography Molecular,Protein Proteins/*metabolism/physiology,Escherichia Proteins/*metabolism/physiology,Models Proteins/metabolism,Cell Proteins/metabolism,membrane Relationship,structure-function,Tandem Sequence/genetics,Binding Signals/genetics,protein Sites/genetics,Carrier Sorting Spectrometry/methods,Tat System/*metabolism/physiology,zero Transport Transport/physiology,Structure-Activity assembly,TatB,TatBC coli coli/metabolism,LC-MS/MS,mass complex,protein complex,to_read,translocation,twin-arginine cross-linker cross-linking,protein export,Protein protein,Membrane space spectrometry trafficking,membrane translocase translocation,Protein translocation,Twin-Arginine-Translocation transport,Membrane}, langid = {english}, month = sep, number = 38, pages = {13902--13914}, pmcid = {PMC6755809}, pmid = {31341014}, timestamp = {2024-05-17T13:01:35.000+0200}, title = {Surface-Exposed Domains of {{TatB}} Involved in the Structural and Functional Assembly of the {{Tat}} Translocase in {{Escherichia}} Coli.}, volume = 294, year = 2019 }