scholarly journals Molecular Modeling of Signal Peptide Recognition by Eukaryotic Sec Complexes

2021 ◽  
Vol 22 (19) ◽  
pp. 10705
Author(s):  
Pratiti Bhadra ◽  
Volkhard Helms
Keyword(s):  
Protein Translocation ◽  
Conformational Dynamics ◽  
Primary Component ◽  
Signal Peptides ◽  
Accessory Proteins ◽  
Simulation Studies ◽  
Peptide Recognition ◽  
Substrate Peptide ◽  
Modelling Studies ◽  
Lateral Gate

Here, we review recent molecular modelling and simulation studies of the Sec translocon, the primary component/channel of protein translocation into the endoplasmic reticulum (ER) and bacterial periplasm, respectively. Our focus is placed on the eukaryotic Sec61, but we also mention modelling studies on prokaryotic SecY since both systems operate in related ways. Cryo-EM structures are now available for different conformational states of the Sec61 complex, ranging from the idle or closed state over an inhibited state with the inhibitor mycolactone bound near the lateral gate, up to a translocating state with bound substrate peptide in the translocation pore. For all these states, computational studies have addressed the conformational dynamics of the translocon with respect to the pore ring, the plug region, and the lateral gate. Also, molecular simulations are addressing mechanistic issues of insertion into the ER membrane vs. translocation into the ER, how signal-peptides are recognised at all in the translocation pore, and how accessory proteins affect the Sec61 conformation in the co- and post-translational pathways.

scholarly journals Conformational entropy of a single peptide controlled under force governs protease recognition and catalysis

2018 ◽  
Vol 115 (45) ◽  
pp. 11525-11530 ◽  
Author(s):  
Marcelo E. Guerin ◽  
Guillaume Stirnemann ◽  
David Giganti
Keyword(s):  
Single Molecule ◽  
Conformational Dynamics ◽  
Conformational Sampling ◽  
Enzymatic Reactions ◽  
Sequence Specificity ◽  
Proteomics Data ◽  
Conformational Entropy ◽  
Substrate Peptide ◽  
The Impact

An immense repertoire of protein chemical modifications catalyzed by enzymes is available as proteomics data. Quantifying the impact of the conformational dynamics of the modified peptide remains challenging to understand the decisive kinetics and amino acid sequence specificity of these enzymatic reactions in vivo, because the target peptide must be disordered to accommodate the specific enzyme-binding site. Here, we were able to control the conformation of a single-molecule peptide chain by applying mechanical force to activate and monitor its specific cleavage by a model protease. We found that the conformational entropy impacts the reaction in two distinct ways. First, the flexibility and accessibility of the substrate peptide greatly increase upon mechanical unfolding. Second, the conformational sampling of the disordered peptide drives the specific recognition, revealing force-dependent reaction kinetics. These results support a mechanism of peptide recognition based on conformational selection from an ensemble that we were able to quantify with a torsional free-energy model. Our approach can be used to predict how entropy affects site-specific modifications of proteins and prompts conformational and mechanical selectivity.

scholarly journals Comparative Analysis of Apicoplast-Targeted Protein Extension Lengths in Apicomplexan Parasites

2015 ◽  
Vol 2015 ◽  
pp. 1-6 ◽  
Author(s):  
Alexandr V. Seliverstov ◽  
Oleg A. Zverkov ◽  
Svetlana N. Istomina ◽  
Sergey A. Pirogov ◽  
Philip S. Kitsis
Keyword(s):  
Plasmodium Falciparum ◽  
Comparative Analysis ◽  
Protein Translocation ◽  
Neospora Caninum ◽  
Signal Peptides ◽  
Model Species ◽  
Apicomplexan Parasites ◽  
Important Region ◽  
Focus On Results ◽  
Species Comparisons

In general, the mechanism of protein translocation through the apicoplast membrane requires a specific extension of a functionally important region of the apicoplast-targeted proteins. The corresponding signal peptides were detected in many apicomplexans but not in the majority of apicoplast-targeted proteins inToxoplasma gondii. InT. gondiisignal peptides are either much diverged or their extension region is processed, which in either case makes the situation different from other studied apicomplexans. We propose a statistic method to compare extensions of the functionally important regions of apicoplast-targeted proteins. More specifically, we provide a comparison of extension lengths of orthologous apicoplast-targeted proteins in apicomplexan parasites. We focus on results obtained for the model speciesT. gondii,Neospora caninum, andPlasmodium falciparum. With our method, cross species comparisons demonstrate that, in average, apicoplast-targeted protein extensions inT. gondiiare 1.5-fold longer than inN. caninumand 2-fold longer than inP. falciparum. Extensions inP. falciparumless than 87 residues in size are longer than the corresponding extensions inN. caninumand, reversely, are shorter if they exceed 88 residues.

scholarly journals Translocation channel gating kinetics balances protein translocation efficiency with signal sequence recognition fidelity

2011 ◽  
Vol 22 (17) ◽  
pp. 2983-2993 ◽  
Author(s):  
Steven F. Trueman ◽  
Elisabet C. Mandon ◽  
Reid Gilmore
Keyword(s):  
Structural Transition ◽  
Protein Translocation ◽  
Signal Sequence ◽  
Channel Gating ◽  
Critical Event ◽  
Protein Insertion ◽  
Sequence Recognition ◽  
Translocation Efficiency ◽  
Lateral Gate ◽  
Insight Into

The transition between the closed and open conformations of the Sec61 complex permits nascent protein insertion into the translocation channel. A critical event in this structural transition is the opening of the lateral translocon gate that is formed by four transmembrane (TM) spans (TM2, TM3, TM7, and TM8 in Sec61p) to expose the signal sequence–binding site. To gain mechanistic insight into lateral gate opening, mutations were introduced into a lumenal loop (L7) that connects TM7 and TM8. The sec61 L7 mutants were found to have defects in both the posttranslational and cotranslational translocation pathways due to a kinetic delay in channel gating. The translocation defect caused by L7 mutations could be suppressed by the prl class of sec61 alleles, which reduce the fidelity of signal sequence recognition. The prl mutants are proposed to act by destabilizing the closed conformation of the translocation channel. Our results indicate that the equilibrium between the open and closed conformations of the protein translocation channel maintains a balance between translocation activity and signal sequence recognition fidelity.

Conformational dynamics of helix S6 from Shaker potassium channel: Simulation studies

Biopolymers ◽  
2002 ◽  
Vol 64 (6) ◽  
pp. 303-313 ◽  
Author(s):  
Joanne N. Bright ◽  
Indira H. Shrivastava ◽  
Frank S. Cordes ◽  
Mark S. P. Sansom
Keyword(s):  
Potassium Channel ◽  
Conformational Dynamics ◽  
Simulation Studies ◽  
Shaker Potassium Channel ◽  
Channel Simulation

scholarly journals Inter-domain dynamics and fibrinolytic activity of matrix metalloprotease-1

2019 ◽  
Author(s):  
Lokender Kumar ◽  
Joan Planas-Iglesias ◽  
Chase Harms ◽  
Sumaer Kamboj ◽  
Derek Wright ◽  
...  
Keyword(s):  
Single Molecule ◽  
Blood Clot ◽  
Conformational Dynamics ◽  
Resonance Energy Transfer ◽  
Resonance Energy ◽  
Primary Component ◽  
Water Soluble ◽  
Matrix Metalloprotease ◽  
Allosteric Control ◽  
Domain Dynamics

AbstractThe roles of protein conformational dynamics and allostery in function are well-known. However, the roles that inter-domain dynamics have in function are not entirely understood. We used matrix metalloprotease-1 (MMP1) as a model system to study the relationship between inter-domain dynamics and activity because MMP1 has diverse substrates. Here we focus on fibrin, the primary component of a blood clot. Water-soluble fibrinogen, following cleavage by thrombin, self-polymerize to form water-insoluble fibrin. We studied the inter-domain dynamics of MMP1 on fibrin without crosslinks using single-molecule Forster Resonance Energy Transfer (smFRET). We observed that the distance between the catalytic and hemopexin domains of MMP1 increases or decreases as the MMP1 activity increases or decreases, respectively. We modulated the activity using 1) an active site mutant (E219Q) of MMP1, 2) MMP9, another member of the MMP family that increases the activity of MMP1, and 3) tetracycline, an inhibitor of MMP1. We fitted the histograms of smFRET values to a sum of two Gaussians and the autocorrelations to an exponential and power law. We modeled the dynamics as a two-state Poisson process and calculated the kinetic rates from the histograms and autocorrelations. Activity-dependent inter-domain dynamics may enable allosteric control of the MMP1 function.

scholarly journals Structure of the posttranslational Sec protein-translocation channel complex from yeast

Science ◽  
2018 ◽  
Vol 363 (6422) ◽  
pp. 84-87 ◽  
Author(s):  
Samuel Itskanov ◽  
Eunyong Park
Keyword(s):  
Electron Microscopy ◽  
Saccharomyces Cerevisiae ◽  
Endoplasmic Reticulum ◽  
Membrane Proteins ◽  
Binding Sites ◽  
Protein Translocation ◽  
Secretory Proteins ◽  
Conducting Channel ◽  
Cryo Electron Microscopy ◽  
Lateral Gate

The Sec61 protein-conducting channel mediates transport of many proteins, such as secretory proteins, across the endoplasmic reticulum (ER) membrane during or after translation. Posttranslational transport is enabled by two additional membrane proteins associated with the channel, Sec63 and Sec62, but its mechanism is poorly understood. We determined a structure of the Sec complex (Sec61-Sec63-Sec71-Sec72) from Saccharomyces cerevisiae by cryo–electron microscopy (cryo-EM). The structure shows that Sec63 tightly associates with Sec61 through interactions in cytosolic, transmembrane, and ER-luminal domains, prying open Sec61’s lateral gate and translocation pore and thus activating the channel for substrate engagement. Furthermore, Sec63 optimally positions binding sites for cytosolic and luminal chaperones in the complex to enable efficient polypeptide translocation. Our study provides mechanistic insights into eukaryotic posttranslational protein translocation.

scholarly journals A signal sequence suppressor mutant that stabilizes an assembled state of the twin arginine translocase

2017 ◽  
Vol 114 (10) ◽  
pp. E1958-E1967 ◽  
Author(s):  
Qi Huang ◽  
Felicity Alcock ◽  
Holger Kneuper ◽  
Justin C. Deme ◽  
Sarah E. Rollauer ◽  
...  
Keyword(s):  
Signal Peptide ◽  
Protein Translocation ◽  
Signal Sequence ◽  
Transmembrane Helix ◽  
Peptide Binding ◽  
Signal Peptides ◽  
Arginine Residues ◽  
Tat System ◽  
Twin Arginine Translocase

The twin-arginine protein translocation (Tat) system mediates transport of folded proteins across the cytoplasmic membrane of bacteria and the thylakoid membrane of chloroplasts. The Tat system ofEscherichia coliis made up of TatA, TatB, and TatC components. TatBC comprise the substrate receptor complex, and active Tat translocases are formed by the substrate-induced association of TatA oligomers with this receptor. Proteins are targeted to TatBC by signal peptides containing an essential pair of arginine residues. We isolated substitutions, locating to the transmembrane helix of TatB that restored transport activity to Tat signal peptides with inactivating twin arginine substitutions. A subset of these variants also suppressed inactivating substitutions in the signal peptide binding site on TatC. The suppressors did not function by restoring detectable signal peptide binding to the TatBC complex. Instead, site-specific cross-linking experiments indicate that the suppressor substitutions induce conformational change in the complex and movement of the TatB subunit. The TatB F13Y substitution was associated with the strongest suppressing activity, even allowing transport of a Tat substrate lacking a signal peptide. In vivo analysis using a TatA–YFP fusion showed that the TatB F13Y substitution resulted in signal peptide-independent assembly of the Tat translocase. We conclude that Tat signal peptides play roles in substrate targeting and in triggering assembly of the active translocase.

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