scholarly journals Energy landscape steering in SecYEG mediates dynamic coupling in ATP driven protein translocation

2019 ◽  
Author(s):  
Joel A. Crossley ◽  
Tomas Fessl ◽  
Matthew A. Watson ◽  
Daniel W. Watkins ◽  
Robin A. Corey ◽  
...  
Keyword(s):  
Lipid Bilayers ◽  
Conformational Changes ◽  
Protein Translocation ◽  
Energy Landscape ◽  
Membrane Pore ◽  
Reaction Coordinates ◽  
Allosteric Communication ◽  
Pore Opening ◽  
Lateral Gate ◽  
Allosteric Model

AbstractThe Sec translocon is a transmembrane assembly highly conserved among all forms of life as the principal route for transport of polypeptides across or into lipid bilayers. In bacteria translocation involves allosteric communication between the membrane pore SecYEG and the associated SecA ATPase. Using singlemolecule fluorescence we reveal that slow conformational changes associated with the ATPase SecA modulate fast opening and closure of the SecY lateral gate. Such a mismatch of timescales is not compatible with direct coupling between SecA and SecYEG. A dynamic allosteric model is proposed in which the SecA ATPase cycle ‘steers’ the energy landscape for SecY pore opening. We map the experimental traces onto reduced reaction coordinates derived from molecular dynamics trajectories, providing a model for the energy landscape and a structural interpretation of the associated dynamics. Dynamic allostery may be common among motor ATPases that drive conformational changes in molecular machines.Graphical TOC Entry

scholarly journals Metastable Intermediates as Stepping Stones on the Maturation Pathways of Viral Capsids

mBio ◽  
2014 ◽  
Vol 5 (6) ◽  
Author(s):  
Giovanni Cardone ◽  
Robert L. Duda ◽  
Naiqian Cheng ◽  
Lili You ◽  
James F. Conway ◽  
...  
Keyword(s):  
Conformational Changes ◽  
Large Scale ◽  
Structural Changes ◽  
Energy Landscape ◽  
Potential Hazard ◽  
Stepping Stones ◽  
Scanning Calorimetry ◽  
Conformational Effects ◽  
Capsid Maturation ◽  
Capsid Integrity

ABSTRACT As they mature, many capsids undergo massive conformational changes that transform their stability, reactivity, and capacity for DNA. In some cases, maturation proceeds via one or more intermediate states. These structures represent local minima in a rich energy landscape that combines contributions from subunit folding, association of subunits into capsomers, and intercapsomer interactions. We have used scanning calorimetry and cryo-electron microscopy to explore the range of capsid conformations accessible to bacteriophage HK97. To separate conformational effects from those associated with covalent cross-linking (a stabilization mechanism of HK97), a cross-link-incompetent mutant was used. The mature capsid Head I undergoes an endothermic phase transition at 60°C in which it shrinks by 7%, primarily through changes in its hexamer conformation. The transition is reversible, with a half-life of ~3 min; however, >50% of reverted capsids are severely distorted or ruptured. This observation implies that such damage is a potential hazard of large-scale structural changes such as those involved in maturation. Assuming that the risk is lower for smaller changes, this suggests a rationalization for the existence of metastable intermediates: that they serve as stepping stones that preserve capsid integrity as it switches between the radically different conformations of its precursor and mature states. IMPORTANCE Large-scale conformational changes are widespread in virus maturation and infection processes. These changes are accompanied by the release of conformational free energy as the virion (or fusogenic glycoprotein) switches from a precursor state to its mature state. Each state corresponds to a local minimum in an energy landscape. The conformational changes in capsid maturation are so radical that the question arises of how maturing capsids avoid being torn apart. Offering proof of principle, severe damage is inflicted when a bacteriophage HK97 capsid reverts from the (nonphysiological) state that it enters when heated past 60°C. We suggest that capsid proteins have been selected in part by the criterion of being able to avoid sustaining collateral damage as they mature. One way of achieving this—as with the HK97 capsid—involves breaking the overall transition down into several smaller steps in which the risk of damage is reduced.

scholarly journals Free energy landscape for the entire transport cycle of triose-phosphate/phosphate translocator

2017 ◽  
Author(s):  
Mizuki Takemoto ◽  
Yongchan Lee ◽  
Ryuichiro Ishitani ◽  
Osamu Nureki
Keyword(s):  
Free Energy ◽  
Conformational Changes ◽  
Conformational Transition ◽  
Substrate Binding ◽  
Energy Landscape ◽  
Umbrella Sampling ◽  
Free Energy Landscape ◽  
Coupling Mechanism ◽  
Triose Phosphate ◽  
Phosphate Translocator

AbstractSecondary active transporters translocate their substrates using the electrochemical potentials of other chemicals, undergoing large-scale conformational changes. Despite extensive structural studies, the atomic details of the transport mechanism still remain elusive. Here we performed a series of all-atom molecular dynamics simulations of the triose-phosphate/phosphate translocator (TPT), which exports organic phosphates in the chloroplast stroma in strict counter exchange with inorganic phosphate (Pi). Biased sampling methods, including string method and umbrella sampling, successfully reproduced the conformational changes between the inward– and outward-facing states, along with the substrate binding. The free energy landscape of this entire TPT transition pathway demonstrated the alternating access and substrate translocation mechanisms, which revealed Pi is relayed by positively charged residues along the transition pathway. Furthermore, the conserved Glu207 functions as a “molecular switch”, linking the local substrate binding and the global conformational transition. Our results provide atomic-detailed insights into the energy coupling mechanism of antiporter.

scholarly journals Contribution of different effluent organic matter fractions to membrane fouling in ultrafiltration of treated domestic wastewater

2012 ◽  
Vol 2 (4) ◽  
pp. 204-209 ◽  
Author(s):  
X. Zheng ◽  
J. P. Croue
Keyword(s):  
Organic Matter ◽  
Membrane Fouling ◽  
Domestic Wastewater ◽  
Molecular Size ◽  
Effluent Organic Matter ◽  
Membrane Pore ◽  
Pore Opening ◽  
Organic Matter Fractions ◽  
Doc Load ◽  
Hydrophobic Acids

In the present work, effluent organic matter (EfOM) in treated domestic wastewater was separated into hydrophobic neutrals, colloids, hydrophobic acids, transphilic acids and neutrals and hydrophilic compounds. Their contribution to dissolved organic carbon (DOC) was identified. Further characterization was conducted with respect to molecular size and hydrophobicity. Each isolated fraction was dosed into salt solution to identify its fouling potential in ultrafiltration (UF) using a hydrophilized polyethersulfone membrane. The results show that each kind of EfOM leads to irreversible fouling. At similar delivered DOC load to the membrane, colloids present the highest fouling effect in terms of both reversible and irreversible fouling. The hydrophobic organics show much lower reversibility than the biopolymers present. However, as they are of much smaller size than the membrane pore opening, they cannot lead to such severe fouling as biopolymers do. In all of the isolated fractions, hydrophilics show the lowest fouling potential. For either colloids or hydrophobic substances, increasing their content in feedwater leads to worse fouling. The co-effect between biopolymers and other EfOM fractions has also been identified as one of the mechanisms contributing to UF fouling in filtering EfOM-containing waters.

scholarly journals Correction to Protein Translocation Activity in Surface-Supported Lipid Bilayers

Langmuir ◽  
2020 ◽  
Vol 36 (22) ◽  
pp. 6323-6323 ◽  
Author(s):  
Kanokporn Chattrakun ◽  
David P. Hoogerheide ◽  
Chunfeng Mao ◽  
Linda L. Randall ◽  
Gavin M. King
Keyword(s):  
Lipid Bilayers ◽  
Protein Translocation ◽  
Supported Lipid Bilayers

scholarly journals Free-energy landscape of molecular interactions between endothelin 1 and human endothelin type B receptor: fly-casting mechanism

2019 ◽  
Vol 32 (7) ◽  
pp. 297-308 ◽  
Author(s):  
Junichi Higo ◽  
Kota Kasahara ◽  
Mitsuhito Wada ◽  
Bhaskar Dasgupta ◽  
Narutoshi Kamiya ◽  
...  
Keyword(s):  
Free Energy ◽  
Energy Landscape ◽  
Binding Pocket ◽  
Free Energy Landscape ◽  
Type B ◽  
Endothelin 1 ◽  
Reaction Coordinates ◽  
Explicit Solvent ◽  
Virtual System ◽  
G Protein Coupled

Abstract The free-energy landscape of interaction between a medium-sized peptide, endothelin 1 (ET1), and its receptor, human endothelin type B receptor (hETB), was computed using multidimensional virtual-system coupled molecular dynamics, which controls the system’s motions by introducing multiple reaction coordinates. The hETB embedded in lipid bilayer was immersed in explicit solvent. All molecules were expressed as all-atom models. The resultant free-energy landscape had five ranges with decreasing ET1–hETB distance: completely dissociative, outside-gate, gate, binding pocket, and genuine-bound ranges. In the completely dissociative range, no ET1–hETB interaction appeared. In the outside-gate range, an ET1–hETB attractive interaction was the fly-casting mechanism. In the gate range, the ET1 orientational variety decreased rapidly. In the binding pocket range, ET1 was in a narrow pathway with a steep free-energy slope. In the genuine-bound range, ET1 was in a stable free-energy basin. A G-protein-coupled receptor (GPCR) might capture its ligand from a distant place.

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.

scholarly journals SecA Dimer Cross-Linked at Its Subunit Interface Is Functional for Protein Translocation

2006 ◽  
Vol 188 (1) ◽  
pp. 335-338 ◽  
Author(s):  
Lucia B. Jilaveanu ◽  
Donald Oliver
Keyword(s):  
Conformational Changes ◽  
Protein Transport ◽  
Protein Translocation ◽  
Directed Mutagenesis ◽  
Subunit Interface ◽  
Cargo Proteins ◽  
Considerable Controversy ◽  
The Subject ◽  
Vitro Protein

ABSTRACT SecA facilitates protein transport across the eubacterial plasma membrane by its association with cargo proteins and the SecYEG translocon, followed by ATP-driven conformational changes that promote protein translocation in a stepwise manner. Whether SecA functions as a monomer or a dimer during this process has been the subject of considerable controversy. Here we utilize cysteine-directed mutagenesis along with the crystal structure of the SecA dimer to create a cross-linked dimer at its subunit interface, which was normally active for in vitro protein translocation.

The structure of aquaporins

2006 ◽  
Vol 39 (4) ◽  
pp. 361-396 ◽  
Author(s):  
Tamir Gonen ◽  
Thomas Walz
Keyword(s):  
Lipid Bilayers ◽  
Conformational Changes ◽  
Hormonal Regulation ◽  
Structural Information ◽  
Current Knowledge ◽  
Structural Features ◽  
E Coli ◽  
Water Pore ◽  
Water Conduction ◽  
Transmembrane Pores

1. Introduction 3621.1 The elusive water pores 3621.2 CHIP28 3622. Studies on AQP-1 3632.1 Expression of AQP1 cDNA in Xenopus oocytes 3632.2 Reconstitution of purified AQP1 into artificial lipid bilayers 3642.3 Structural information deduced from the primary sequence 3652.4 Evolution and mammalian AQPs 3653. Chronological overview over AQP structures 3683.1 AQP1 – the red blood cell water pore 3683.2 GlpF – the E. coli glycerol facilitator 3713.3 AQPZ – the E. coli water pore 3723.4 AQP0 – the lens-specific aquaporin 3733.5 AQP4 – the main aquaporin in brain 3773.6 SoPiP2;1 – a plant aquaporin 3793.7 AQPM – an archaeabacterial aquaporin 3794. Proton exclusion 3805. Substrate selectivity 3826. Pore regulation 3856.1 Hormonal regulation of AQP trafficking 3856.2 Influence of pH on AQP water conduction 3866.3 Regulation of AQP pore conductance by protein binding 3876.4 Pore closure by conformational changes in the AQP0 pore 3887. Unresolved questions 3908. Acknowledgments 3909. References 391The ubiquitous members of the aquaporin (AQP) family form transmembrane pores that are either exclusive for water (aquaporins) or are also permeable for other small neutral solutes such as glycerol (aquaglyceroporins). The purpose of this review is to provide an overview of our current knowledge of AQP structures and to describe the structural features that define the function of these membrane pores. The review will discuss the mechanisms governing water conduction, proton exclusion and substrate specificity, and how the pore permeability is regulated in different members of the AQP family.

scholarly journals The Mitochondrial Oxidase Assembly Protein1 (Oxa1) Insertase Forms a Membrane Pore in Lipid Bilayers

2012 ◽  
Vol 287 (40) ◽  
pp. 33314-33326 ◽  
Author(s):  
Vivien Krüger ◽  
Markus Deckers ◽  
Markus Hildenbeutel ◽  
Martin van der Laan ◽  
Maike Hellmers ◽  
...  
Keyword(s):  
Lipid Bilayers ◽  
Membrane Pore

scholarly journals Additional In Vitro and In Vivo Evidence for SecA Functioning as Dimers in the Membrane: Dissociation into Monomers Is Not Essential for Protein Translocation in Escherichia coli

2007 ◽  
Vol 190 (4) ◽  
pp. 1413-1418 ◽  
Author(s):  
Hongyun Wang ◽  
Bing Na ◽  
Hsiuchin Yang ◽  
Phang C. Tai
Keyword(s):  
Escherichia Coli ◽  
Lipid Bilayers ◽  
Cytoplasmic Membrane ◽  
Protein Translocation ◽  
Secretory Proteins ◽  
Oligomeric State ◽  
Dependent Protein ◽  
Ts Mutant

ABSTRACT SecA is an essential component in the Sec-dependent protein translocation pathway and, together with ATP, provides the driving force for the transport of secretory proteins across the cytoplasmic membrane of Escherichia coli. Previous studies established that SecA undergoes monomer-dimer equilibrium in solution. However, the oligomeric state of functional SecA during the protein translocation process is controversial. In this study, we provide additional evidence that SecA functions as a dimer in the membrane by (i) demonstration of the capability of the presumably monomeric SecA derivative to be cross-linked as dimers in vitro and in vivo, (ii) complementation of the growth of a secA(Ts) mutant with another nonfunctional SecA or (iii) in vivo complementation and in vitro function of a genetically tandem SecA dimer that does not dissociate into monomers, and (iv) formation of similar ring-like structures by the tandem SecA dimer and SecA in the presence of lipid bilayers. We conclude that SecA functions as a dimer in the membrane and dissociation into monomers is not necessary during protein translocation.

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