scholarly journals The metastable XBP1u transmembrane domain defines determinants for intramembrane proteolysis by signal peptide peptidase

2018 ◽  
Author(s):  
Sara Suna Yücel ◽  
Walter Stelzer ◽  
Alessandra Lorenzoni ◽  
Manfred Wozny ◽  
Dieter Langosch ◽  
...  
Keyword(s):  
Signal Peptide ◽  
Transmembrane Domain ◽  
Chain Complex ◽  
Peptide Bond ◽  
Intramembrane Proteolysis ◽  
Signal Peptide Peptidase ◽  
Cellular Assays ◽  
Nascent Chain ◽  
The Unfolded Protein Response ◽  
The Impact

SummaryUnspliced XBP1 mRNA encodes XBP1u, the transcriptionally inert variant of the unfolded protein response (UPR) transcription factor XBP1s. XBP1u targets its mRNA-ribosome-nascent-chain-complex to the endoplasmic reticulum (ER) to facilitate UPR activation and prevents overactivation. Yet, its membrane association is controversial. Here, we use cell-free translocation and cellular assays to define a moderately hydrophobic stretch in XBP1u that is sufficient to mediate insertion into the ER membrane. Mutagenesis of this transmembrane (TM) region reveals residues that facilitate XBP1u turnover by an ER-associated degradation route that is dependent on signal peptide peptidase (SPP). Furthermore, the impact of these mutations on TM helix dynamics was assessed by residue-specific amide exchange kinetics, evaluated by a semi-automated algorithm. Based on our results, we suggest that SPP-catalyzed intramembrane proteolysis of TM helices is not only determined by their conformational flexibility, but also by side chain interactions near the scissile peptide bond with the enzyme’s active site.

scholarly journals The Metastable XBP1u Transmembrane Domain Defines Determinants for Intramembrane Proteolysis by Signal Peptide Peptidase

Cell Reports ◽  
2019 ◽  
Vol 26 (11) ◽  
pp. 3087-3099.e11 ◽  
Author(s):  
Sara Suna Yücel ◽  
Walter Stelzer ◽  
Alessandra Lorenzoni ◽  
Manfred Wozny ◽  
Dieter Langosch ◽  
...  
Keyword(s):  
Signal Peptide ◽  
Transmembrane Domain ◽  
Intramembrane Proteolysis ◽  
Signal Peptide Peptidase

scholarly journals Signal peptide peptidase activity connects the unfolded protein response to plant defense suppression by Ustilago maydis

2019 ◽  
Vol 15 (4) ◽  
pp. e1007734 ◽  
Author(s):  
Niko Pinter ◽  
Christina Andrea Hach ◽  
Martin Hampel ◽  
Dmitrij Rekhter ◽  
Krzysztof Zienkiewicz ◽  
...  
Keyword(s):  
Plant Defense ◽  
Unfolded Protein Response ◽  
Signal Peptide ◽  
Ustilago Maydis ◽  
Peptidase Activity ◽  
Signal Peptide Peptidase ◽  
Unfolded Protein ◽  
The Unfolded Protein Response ◽  
Protein Response

scholarly journals Biochemical Properties of a Putative Signal Peptide Peptidase from the Hyperthermophilic Archaeon Thermococcus kodakaraensis KOD1

2005 ◽  
Vol 187 (20) ◽  
pp. 7072-7080 ◽  
Author(s):  
Rie Matsumi ◽  
Haruyuki Atomi ◽  
Tadayuki Imanaka
Keyword(s):  
Substrate Specificity ◽  
Signal Peptide ◽  
Biochemical Characterization ◽  
Transmembrane Domain ◽  
Biochemical Properties ◽  
Peptidase Activity ◽  
Amino Acid Residues ◽  
Hyperthermophilic Archaeon ◽  
Signal Peptide Peptidase ◽  
Putative Signal Peptide

ABSTRACT We have performed the first biochemical characterization of a putative archaeal signal peptide peptidase (SppATk) from the hyperthermophilic archaeon Thermococcus kodakaraensis KOD1. SppATk, comprised of 334 residues, was much smaller than its counterpart from Escherichia coli (618 residues) and harbored a single predicted transmembrane domain near its N terminus. A truncated mutant protein without the N-terminal 54 amino acid residues (ΔN54SppATk) was found to be stable against autoproteolysis and was examined further. ΔN54SppATk exhibited peptidase activity towards fluorogenic peptide substrates and was found to be highly thermostable. Moreover, the enzyme displayed a remarkable stability and preference for alkaline pH, with optimal activity detected at pH 10. ΔN54SppATk displayed a Km of 240 ± 18 μM and a V max of 27.8 ± 0.7 μmol min−1 mg−1 towards Ala-Ala-Phe-4-methyl-coumaryl-7-amide at 80°C and pH 10. The substrate specificity of the enzyme was examined in detail with a FRETS peptide library. By analyzing the cleavage products with liquid chromatography-mass spectrometry, ΔN54SppATk was found to efficiently cleave peptides with a relatively small side chain at the P-1 position and a hydrophobic or aromatic residue at the P-3 position. The positively charged Arg residue was preferred at the P-4 position, while substrates with negatively charged residues at the P-2, P-3, or P-4 position were not cleaved. When predicted signal sequences from the T. kodakaraensis genome sequence were examined, we found that the substrate specificity of ΔN54SppATk was in good agreement with its presumed role as a signal peptide peptidase in this archaeon.

scholarly journals Early encounters of a nascent membrane protein

2005 ◽  
Vol 170 (1) ◽  
pp. 27-35 ◽  
Author(s):  
Edith N.G. Houben ◽  
Raz Zarivach ◽  
Bauke Oudega ◽  
Joen Luirink
Keyword(s):  
Membrane Protein ◽  
Ribosomal Proteins ◽  
Transmembrane Domain ◽  
Signal Recognition Particle ◽  
Chain Complex ◽  
Signal Recognition ◽  
Inner Membrane ◽  
Ribosomal Tunnel ◽  
Nascent Chain ◽  
Inner Membrane Protein

An unbiased photo–cross-linking approach was used to probe the “molecular path” of a growing nascent Escherichia coli inner membrane protein (IMP) from the peptidyl transferase center to the surface of the ribosome. The nascent chain was initially in proximity to the ribosomal proteins L4 and L22 and subsequently contacted L23, which is indicative of progression through the ribosome via the main ribosomal tunnel. The signal recognition particle (SRP) started to interact with the nascent IMP and to target the ribosome–nascent chain complex to the Sec–YidC complex in the inner membrane when maximally half of the transmembrane domain (TM) was exposed from the ribosomal exit. The combined data suggest a flexible tunnel that may accommodate partially folded nascent proteins and parts of the SRP and SecY. Intraribosomal contacts of the nascent chain were not influenced by the presence of a functional TM in the ribosome.

scholarly journals The α-Helical Content of the Transmembrane Domain of the British Dementia Protein-2 (Bri2) Determines Its Processing by Signal Peptide Peptidase-like 2b (SPPL2b)

2011 ◽  
Vol 287 (7) ◽  
pp. 5156-5163 ◽  
Author(s):  
Regina Fluhrer ◽  
Lucas Martin ◽  
Bärbel Klier ◽  
Martina Haug-Kröper ◽  
Gudula Grammer ◽  
...  
Keyword(s):  
Signal Peptide ◽  
Transmembrane Domain ◽  
Signal Peptide Peptidase ◽  
Helical Content

scholarly journals Signal-peptide-peptidase-like 2a is required for CD74 intramembrane proteolysis in human B cells

2014 ◽  
Vol 451 (1) ◽  
pp. 48-53 ◽  
Author(s):  
Janna Schneppenheim ◽  
Susann Hüttl ◽  
Anne Kruchen ◽  
Regina Fluhrer ◽  
Ingo Müller ◽  
...  
Keyword(s):  
B Cells ◽  
Signal Peptide ◽  
Intramembrane Proteolysis ◽  
Signal Peptide Peptidase ◽  
Human B Cells
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