scholarly journals Substrate Binding Drives Large-Scale Conformational Changes in the Hsp90 Molecular Chaperone

2011 ◽  
Vol 42 (1) ◽  
pp. 96-105 ◽  
Author(s):  
Timothy O. Street ◽  
Laura A. Lavery ◽  
David A. Agard
Keyword(s):  
Conformational Changes ◽  
Molecular Chaperone ◽  
Large Scale ◽  
Substrate Binding

scholarly journals Structural polymorphism and substrate promiscuity of a ribosome-associated molecular chaperone

2021 ◽  
Vol 2 (1) ◽  
pp. 375-386
Author(s):  
Chih-Ting Huang ◽  
Yei-Chen Lai ◽  
Szu-Yun Chen ◽  
Meng-Ru Ho ◽  
Yun-Wei Chiang ◽  
...  
Keyword(s):  
Conformational Changes ◽  
Molecular Chaperone ◽  
Binding Sites ◽  
Substrate Binding ◽  
Chaperone Activity ◽  
Trigger Factor ◽  
Peptide Fragments ◽  
Structural Polymorphism ◽  
Ribosome Binding ◽  
Multiple Binding Sites

Abstract. Trigger factor (TF) is a highly conserved multi-domain molecular chaperone that exerts its chaperone activity at the ribosomal tunnel exit from which newly synthesized nascent chains emerge. TF also displays promiscuous substrate binding for a large number of cytosolic proteins independent of ribosome binding. We asked how TF recognizes a variety of substrates while existing in a monomer–dimer equilibrium. Paramagnetic nuclear magnetic resonance (NMR) and electron spin resonance (ESR) spectroscopy were used to show that dimeric TF displays a high degree of structural polymorphism in solution. A series of peptides has been generated to quantify their TF binding affinities in relation with their sequence compositions. The results confirmed a previous predication that TF preferentially binds to peptide fragments that are rich in aromatic and positively charged amino acids. NMR paramagnetic relaxation enhancement analysis showed that TF utilizes multiple binding sites, located in the chaperone domain and part of the prolyl trans–cis isomerization domain, to interact with these peptides. Dimerization of TF effectively sequesters most of the substrate binding sites, which are expected to become accessible upon binding to the ribosome as a monomer. As TF lacks ATPase activity, which is commonly used to trigger conformational changes within molecular chaperones in action, the ribosome-binding-associated disassembly and conformational rearrangements may be the underlying regulatory mechanism of its chaperone activity.

scholarly journals Structural polymorphism and substrate promiscuity of a ribosome-associated molecular chaperone

2021 ◽  
Author(s):  
Chih-Ting Huang ◽  
Yei-Chen Lai ◽  
Szu-Yun Chen ◽  
Meng-Ru Ho ◽  
Yun-Wei Chiang ◽  
...  
Keyword(s):  
Conformational Changes ◽  
Molecular Chaperone ◽  
Binding Sites ◽  
Substrate Binding ◽  
Chaperone Activity ◽  
Trigger Factor ◽  
Paramagnetic Nmr ◽  
Resonance Spectroscopy ◽  
Structural Polymorphism ◽  
Ribosome Binding

Abstract. Trigger factor (TF) is a highly conserved multi-domain molecular chaperone that exerts its chaperone activity at the ribosomal tunnel exit from which newly synthesized nascent chains emerge. TF also displays promiscuous substrate binding for a large number of cytosolic proteins independent of ribosome binding. We asked how TF recognizes a variety of substrates while existing in a monomer-dimer equilibrium. Paramagnetic NMR, electron spin resonance spectroscopy and chemical crosslink show that dimeric TF displays a high degree of structural polymorphism in solution. A series of peptides has been generated to quantify their TF binding affinities in relation with their sequence compositions. The results confirmed a previous predication that TF preferentially binds to peptide fragments that are rich in aromatic and positively charged amino acids. NMR paramagnetic relaxation enhancement analysis showed that TF utilizes multiple binding sites, located in the chaperone domain and part of the prolyl trans/cis isomerisation domain, to interact with these peptides. Dimerization of TF effectively sequesters most of substrate binding sites, which are expected to become accessible upon binding to the ribosome as a monomer. As TF lacks ATPase activity, which is commonly used to trigger conformational changes within molecular chaperones in action, the ribosome-binding-associated disassembly and conformational rearrangements may be the underlying regulatory mechanism of its chaperone activity.

Insights into the roles of charged residues in substrate binding and mode of action of mannuronan C-5 epimerase AlgE4

Glycobiology ◽  
2021 ◽  
Author(s):  
Margrethe Gaardløs ◽  
Sergey A Samsonov ◽  
Marit Sletmoen ◽  
Maya Hjørnevik ◽  
Gerd Inger Sætrom ◽  
...  
Keyword(s):  
Optical Tweezers ◽  
Conformational Changes ◽  
Molecular Mechanisms ◽  
Hydrophobic Interactions ◽  
Substrate Binding ◽  
Interdisciplinary Approach ◽  
Product Formation ◽  
Titration Calorimetry ◽  
Binding Groove ◽  
Dynamics Simulations

Abstract Mannuronan C-5 epimerases catalyse the epimerization of monomer residues in the polysaccharide alginate, changing the physical properties of the biopolymer. The enzymes are utilized to tailor alginate to numerous biological functions by alginate-producing organisms. The underlying molecular mechanisms that control the processive movement of the epimerase along the substrate chain is still elusive. To study this, we have used an interdisciplinary approach combining molecular dynamics simulations with experimental methods from mutant studies of AlgE4, where initial epimerase activity and product formation were addressed with NMR spectroscopy, and characteristics of enzyme-substrate interactions were obtained with isothermal titration calorimetry and optical tweezers. Positive charges lining the substrate-binding groove of AlgE4 appear to control the initial binding of poly-mannuronate, and binding also seems to be mediated by both electrostatic and hydrophobic interactions. After the catalytic reaction, negatively charged enzyme residues might facilitate dissociation of alginate from the positive residues, working like electrostatic switches, allowing the substrate to translocate in the binding groove. Molecular simulations show translocation increments of two monosaccharide units before the next productive binding event resulting in MG-block formation, with the epimerase moving with its N-terminus towards the reducing end of the alginate chain. Our results indicate that the charge pair R343-D345 might be directly involved in conformational changes of a loop that can be important for binding and dissociation. The computational and experimental approaches used in this study complement each other, allowing for a better understanding of individual residues’ roles in binding and movement along the alginate chains.

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 Nuclear pores constrict upon energy depletion

2020 ◽  
Author(s):  
Christian E Zimmerli ◽  
Matteo Allegretti ◽  
Vasileios Rantos ◽  
Sara K Goetz ◽  
Agnieszka Obarska-Kosinska ◽  
...  
Keyword(s):  
Conformational Changes ◽  
Large Scale ◽  
Electron Tomography ◽  
Nuclear Pore ◽  
Nuclear Pore Complexes ◽  
Energy Status ◽  
Energy Depletion ◽  
Time Variations ◽  
Subtomogram Averaging ◽  
Pore Complexes

Nuclear pore complexes (NPCs) fuse the inner and outer nuclear membranes and mediate nucleocytoplasmic exchange. They are made of 30 different nucleoporins that form an intricate cylindrical architecture around an aqueous central channel. This architecture is highly dynamic in space and time. Variations in NPC diameter were reported, but the physiological circumstances and the molecular details remain unknown. Here we combined cryo-electron tomography and subtomogram averaging with integrative structural modeling to capture a molecular movie of the respective large-scale conformational changes in cellulo. While actively transporting NPCs adopt a dilated conformation, they strongly constrict upon cellular energy depletion. Fluorescence recovery after photo bleaching experiments show that NPC constriction is concomitant with reduced diffusion and active transport across the nuclear envelope. Our data point to a model where the energy status of cells is linked to the conformation of NPC architecture.

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 Structural insights into the activation of human calcium-sensing receptor

2021 ◽  
Author(s):  
Xiaochen Chen ◽  
Lu Wang ◽  
Zhanyu Ding ◽  
Qianqian Cui ◽  
Li Han ◽  
...  
Keyword(s):  
Conformational Changes ◽  
Large Scale ◽  
Transmembrane Domains ◽  
Structural Basis ◽  
Calcium Sensing Receptor ◽  
Calcium Sensing ◽  
Cryo Electron Microscopy ◽  
Activation Mechanisms ◽  
G Protein Coupled ◽  
Structural Insights

AbstractHuman calcium-sensing receptor (CaSR) is a G-protein-coupled receptor that maintains Ca2+ homeostasis in serum. Here, we present the cryo-electron microscopy structures of the CaSR in the inactive and active states. Complemented with previously reported crystal structures of CaSR extracellular domains, it suggests that there are three distinct conformations: inactive, intermediate and active state during the activation. We used a negative allosteric nanobody to stabilize the CaSR in the fully inactive state and found a new binding site for Ca2+ ion that acts as a composite agonist with L-amino acid to stabilize the closure of active Venus flytraps. Our data shows that the agonist binding leads to the compaction of the dimer, the proximity of the cysteine-rich domains, the large-scale transitions of 7-transmembrane domains, and the inter-and intrasubunit conformational changes of 7-transmembrane domains to accommodate the downstream transducers. Our results reveal the structural basis for activation mechanisms of the CaSR.

scholarly journals Antigen Presentation of mRNA-Based and Virus-Vectored SARS-CoV-2 Vaccines

Vaccines ◽  
2021 ◽  
Vol 9 (8) ◽  
pp. 848
Author(s):  
Ger T. Rijkers ◽  
Nynke Weterings ◽  
Andres Obregon-Henao ◽  
Michaëla Lepolder ◽  
Taru S. Dutt ◽  
...  
Keyword(s):  
Antigen Presentation ◽  
Conformational Changes ◽  
Dna Sequences ◽  
Neutralizing Antibodies ◽  
Large Scale ◽  
Viral Vector ◽  
Adenovirus Vector ◽  
Platelet Factor ◽  
Immune Mediated ◽  
Protein Encoding

Infection with Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) causes Coronavirus Disease 2019 (COVID-19), which has reached pandemic proportions. A number of effective vaccines have been produced, including mRNA vaccines and viral vector vaccines, which are now being implemented on a large scale in order to control the pandemic. The mRNA vaccines are composed of viral Spike S1 protein encoding mRNA incorporated in a lipid nanoparticle and stabilized by polyethylene glycol (PEG). The mRNA vaccines are novel in many respects, including cellular uptake and the intracellular routing, processing, and secretion of the viral protein. Viral vector vaccines have incorporated DNA sequences, encoding the SARS-CoV-2 Spike protein into (attenuated) adenoviruses. The antigen presentation routes in MHC class I and class II, in relation to the induction of virus-neutralizing antibodies and cytotoxic T-lymphocytes, will be reviewed. In rare cases, mRNA vaccines induce unwanted immune mediated side effects. The mRNA-based vaccines may lead to an anaphylactic reaction. This reaction may be triggered by PEG. The intracellular routing of PEG and potential presentation in the context of CD1 will be discussed. Adenovirus vector-based vaccines have been associated with thrombocytopenic thrombosis events. The anti-platelet factor 4 antibodies found in these patients could be generated due to conformational changes of relevant epitopes presented to the immune system.

scholarly journals Simplified geometric representations of protein structures identify complementary interaction interfaces

2019 ◽  
Author(s):  
Caitlyn L. McCafferty ◽  
Edward M. Marcotte ◽  
David W. Taylor
Keyword(s):  
Conformational Changes ◽  
Protein Interaction ◽  
Protein Function ◽  
Large Scale ◽  
Predictive Accuracy ◽  
Protein Complexes ◽  
Protein Structures ◽  
Molecular Properties ◽  
3D Structures ◽  
Geometric Representations

ABSTRACTProtein-protein interactions are critical to protein function, but three-dimensional (3D) arrangements of interacting proteins have proven hard to predict, even given the identities and 3D structures of the interacting partners. Specifically, identifying the relevant pairwise interaction surfaces remains difficult, often relying on shape complementarity with molecular docking while accounting for molecular motions to optimize rigid 3D translations and rotations. However, such approaches can be computationally expensive, and faster, less accurate approximations may prove useful for large-scale prediction and assembly of 3D structures of multi-protein complexes. We asked if a reduced representation of protein geometry retains enough information about molecular properties to predict pairwise protein interaction interfaces that are tolerant of limited structural rearrangements. Here, we describe a cuboid transformation of 3D protein accessible surfaces on which molecular properties such as charge, hydrophobicity, and mutation rate can be easily mapped, implemented in the MorphProt package. Pairs of surfaces are compared to rapidly assess partner-specific potential surface complementarity. On two available benchmarks of 85 overall known protein complexes, we observed F1 scores (a weighted combination of precision and recall) of 19-34% at correctly identifying protein interaction surfaces, comparable to more computationally intensive 3D docking methods in the annual Critical Assessment of PRedicted Interactions. Furthermore, we examined the effect of molecular motion through normal mode simulation on a benchmark receptor-ligand pair and observed no marked loss of predictive accuracy for distortions of up to 6 Å RMSD. Thus, a cuboid transformation of protein surfaces retains considerable information about surface complementarity, offers enhanced speed of comparison relative to more complex geometric representations, and exhibits tolerance to conformational changes.

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