Membrane Targeting and Insertion of the C-Tail Protein SciP

Soussoula, Lavinia; Seitl, Ines; Lupo, Domenico; Kühn, Andreas

doi:10.1016/j.jmb.2016.09.001

Cited by 30 publications

(23 citation statements)

References 27 publications

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“…It should be noted that insertion of Pf3-Lep and SciP was not as efficient as previously seen 22 , 23 since in the present study both the YidC mutant and the substrate protein are expressed on IPTG inducible plasmids for a short induction time while in the latter studies YidC was continuously expressed at the normal level.…”

Section: Resultscontrasting

confidence: 45%

“…As previously shown, YidC is required for insertion of the C-tailed membrane protein SciP 21 , 22 . The translocation of the small C-tail can be monitored by its modification with 4-acetoamido-4′-maleimidylstilbene-2,2′disodium sulfonate (AMS), where the periplasmic exposure of a cysteine results in an increased molecular weight of 0.5 kDa.…”

Section: Resultsmentioning

confidence: 68%

“…For complementation assays, MK6 was transformed with pGZ119EH 31 expressing the respective YidC proteins under the control of the tac promoter. Coexpression of M13 procoat was performed with pMS-M13procoat 32 and Sci-P with pMS-SciP-C 22 . Where indicated, the M13 procoat wildtype and H5-33C mutant were expressed from the pMS119EH vector.…”

Section: Methodsmentioning

confidence: 99%

“…The plasmid-encoded SciP-C 22 was co-expressed with one of the various defective YidC mutants in MK6 cells in LB (100 µg/mL ampicillin, 37 µg/ml chloramphenicol, 0.4% glucose) under YidC depletion conditions to an OD 600 = 0.5 at 37 °C. After resuspension in M9 medium lacking methionine the cells were grown for another 45 min, induced with 1 mM IPTG for 10 min.…”

Section: Methodsmentioning

confidence: 99%

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Each protomer of a dimeric YidC functions as a single membrane insertase

Spann

Chen

Dalbey

et al. 2018

Sci Rep

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The membrane insertase YidC catalyzes the entrance of newly synthesized proteins into the lipid bilayer. As an integral membrane protein itself, YidC can be found as a monomer, a dimer or also as a member of the holotranslocase SecYEGDF-YajC-YidC. To investigate whether the dimeric YidC is functional and whether two copies cooperate to insert a single substrate, we constructed a fusion protein where two copies of YidC are connected by a short linker peptide. The 120 kDa protein is stable and functional as it supports the membrane insertion of the M13 procoat protein, the C-tailed protein SciP and the fusion protein Pf3-Lep. Mutations that inhibit either protomer do not inactivate the insertase and rather keep it functional. When both protomers are defective, the substrate proteins accumulate in the cytoplasm. This suggests that the dimeric YidC operates as two insertases. Consistent with this, we show that the dimeric YidC can bind two substrate proteins simultaneously, suggesting that YidC indeed functions as a monomer.

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Section: Resultscontrasting

confidence: 45%

Section: Resultsmentioning

confidence: 68%

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

See 2 more Smart Citations

Each protomer of a dimeric YidC functions as a single membrane insertase

Spann

Chen

Dalbey

et al. 2018

Sci Rep

Self Cite

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“…Orientation was tested by PCR and sequencing. The ORF of sfGFP with C-terminal hexa-His tag was cut out from pMS119EH-sfGFP (Pross et al, 2016) with BamHI and HindIII, and subcloned into the BamHI and XbaI sites of pART7, in translational fusion with the propeptide ORFs (HindIII and XbaI sites were blunted). For ER retention, the ORF of sfGFP was extended by PCR to include the C-terminal KDEL sequence, and the PCR product replaced the sfGFP-hexa-His ORF in pART7.…”

Section: Generation Of Expression Constructsmentioning

confidence: 99%

The biogenesis of CLEL peptides involves several processing events in consecutive compartments of the secretory pathway

Stührwohldt

Scholl

Lang

et al. 2018

Preprint

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AbstractPost-translationally modified peptides are involved in many aspects of plant growth and development. The maturation of these peptides from their larger precursors is still poorly understood. We show here that the biogenesis of CLEL6 and CLEL9 peptides in Arabidopsis thaliana requires a series of processing events in consecutive compartments of the secretory pathway. Following cleavage of the signal peptide upon entry into the endoplasmic reticulum (ER), the peptide precursors are processed in the cis-Golgi by the subtilase SBT6.1. SBT6.1-mediated cleavage within the variable domain allows for continued passage of the partially processed precursors through the secretory pathway, and is a prerequisite for subsequent post-translational modifications including tyrosine sulfation and proline hydroxylation within, and proteolytic maturation after exit from the Golgi. Activation by subtilase SBT3.8 in post-Golgi compartments depends on the N-terminal aspartate of the mature peptides. Our work highlights the complexity of post-translational precursor maturation allowing for stringent control of peptide biogenesis.

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Evolutionarily conserved mechanism for membrane recognition from bacteria to mitochondria

2021

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The mechanisms controlling membrane recognition by proteins with one hydrophobic stretch at their carboxyl terminus (tail anchor, TA) are poorly defined. The Escherichia coli TAs of ElaB and YqjD, which share sequential and structural similarity with the Saccharomyces cerevisiae TA of Fis1, were shown to localize to mitochondria. We show that YqjD and ElaB are directed by their TAs to bacterial cell poles. Fis1(TA) expressed in E. coli localizes like the endogenous TAs. The yeast and bacterial TAs are inserted in the E. coli inner membrane, and they all show affiliation to phosphatidic acid (PA), found in the membrane of the bacterial cell poles and of the yeast mitochondria. Our results suggest a mechanism for TA membrane recognition conserved from bacteria to mitochondria and raise the possibility that through their interaction with PA, and TAs play a role across prokaryotes and eukaryotes in controlling cell/organelle fate.

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Membrane Targeting and Insertion of the C-Tail Protein SciP

Cited by 30 publications

References 27 publications

Each protomer of a dimeric YidC functions as a single membrane insertase

Each protomer of a dimeric YidC functions as a single membrane insertase

The biogenesis of CLEL peptides involves several processing events in consecutive compartments of the secretory pathway

Evolutionarily conserved mechanism for membrane recognition from bacteria to mitochondria

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