Molecular Crowding Stabilizes Both the Intrinsically Disordered Calcium-Free State and the Folded Calcium-Bound State of a Repeat in Toxin (RTX) Protein

2013 ◽  
Vol 135 (32) ◽  
pp. 11929-11934 ◽  
Author(s):  
Ana-Cristina Sotomayor-Pérez ◽  
Orso Subrini ◽  
Audrey Hessel ◽  
Daniel Ladant ◽  
Alexandre Chenal
Keyword(s):  
Bound State ◽  
Free State ◽  
Molecular Crowding ◽  
Intrinsically Disordered

scholarly journals Molecular Crowding Stabilizes Both the Intrinsically Disordered Calcium-Free State and the Folded Calcium-Bound State of an RTX Protein: Implication for Toxin Secretion

2014 ◽  
Vol 106 (2) ◽  
pp. 271a
Author(s):  
Ana Cristina Sotomayor Pérez ◽  
Orso Subrini ◽  
Audrey Hessel ◽  
Daniel Ladant ◽  
Alexandre Chenal
Keyword(s):  
Bound State ◽  
Free State ◽  
Molecular Crowding ◽  
Intrinsically Disordered ◽  
Toxin Secretion

scholarly journals Conformational Ensembles of an Intrinsically Disordered Protein pKID with and without a KIX Domain in Explicit Solvent Investigated by All-Atom Multicanonical Molecular Dynamics

Biomolecules ◽  
2012 ◽  
Vol 2 (1) ◽  
pp. 104-121 ◽  
Author(s):  
Koji Umezawa ◽  
Jinzen Ikebe ◽  
Mitsunori Takano ◽  
Haruki Nakamura ◽  
Junichi Higo
Keyword(s):  
Molecular Dynamics ◽  
Free Energy ◽  
Binding Site ◽  
Complex Structure ◽  
Intrinsically Disordered Protein ◽  
Bound State ◽  
Conformational Ensembles ◽  
Intrinsically Disordered ◽  
Dynamics Simulations ◽  
High Flexibility

The phosphorylated kinase-inducible activation domain (pKID) adopts a helix–loop–helix structure upon binding to its partner KIX, although it is unstructured in the unbound state. The N-terminal and C-terminal regions of pKID, which adopt helices in the complex, are called, respectively, αA and αB. We performed all-atom multicanonical molecular dynamics simulations of pKID with and without KIX in explicit solvents to generate conformational ensembles. Although the unbound pKID was disordered overall, αA and αB exhibited a nascent helix propensity; the propensity of αA was stronger than that of αB, which agrees with experimental results. In the bound state, the free-energy landscape of αB involved two low free-energy fractions: native-like and non-native fractions. This result suggests that αB folds according to the induced-fit mechanism. The αB-helix direction was well aligned as in the NMR complex structure, although the αA helix exhibited high flexibility. These results also agree quantitatively with experimental observations. We have detected that the αB helix can bind to another site of KIX, to which another protein MLL also binds with the adopting helix. Consequently, MLL can facilitate pKID binding to the pKID-binding site by blocking the MLL-binding site. This also supports experimentally obtained results.

scholarly journals Multiple proteolytic events in caspase-6 self-activation impact conformations of discrete structural regions

2017 ◽  
Vol 114 (38) ◽  
pp. E7977-E7986 ◽  
Author(s):  
Kevin B. Dagbay ◽  
Jeanne A. Hardy
Keyword(s):  
Substrate Binding ◽  
Complete Removal ◽  
Bound State ◽  
Solvent Exposure ◽  
Strategic Design ◽  
Intrinsically Disordered ◽  
Parkinson’S Diseases ◽  
Active Substrate ◽  
Hydrogen Deuterium ◽  
Caspase 6

Caspase-6 is critical to the neurodegenerative pathways of Alzheimer’s, Huntington’s, and Parkinson’s diseases and has been identified as a potential molecular target for treatment of neurodegeneration. Thus, understanding the global and regional changes in dynamics and conformation provides insights into the unique properties of caspase-6 that may contribute to achieving control of its function. In this work, hydrogen/deuterium exchange MS (H/DX–MS) was used to map the local changes in the conformational flexibility of procaspase-6 at the discrete states that reflect the series of cleavage events that ultimately lead to the fully active, substrate-bound state. Intramolecular self-cleavage at Asp-193 evoked higher solvent exposure in the regions of the substrate-binding loops L1, L3, and L4 and in the 130s region, the intersubunit linker region, the 26–32 region as well as in the stabilized loop 2. Additional removal of the linker allowed caspase-6 to gain more flexibility in the 130s region and in the L2 region converting caspase-6 to a competent substrate-binding state. The prodomain region was found to be intrinsically disordered independent of the activation state of caspase-6; however, its complete removal resulted in the protection of the adjacent 26–32 region, suggesting that this region may play a regulatory role. The molecular details of caspase-6 dynamics in solution provide a comprehensive scaffold for strategic design of therapeutic approaches for neurodegenerative disorders.

scholarly journals Continuous absorption

1930 ◽  
Vol 229 (670-680) ◽  
pp. 163-204 ◽  
Keyword(s):  
Absorption Coefficient ◽  
Bound State ◽  
Free State ◽  
X Rays ◽  
Classical Electromagnetism ◽  
Considerable Difficulty ◽  
Positive Nucleus ◽  
Continuous Absorption ◽  
The Matrix ◽  
Good Agreement

For some years astrophysicists have been looking for an adequate theory of continuous—as opposed to line—absorption. The natural and generally accepted mechanism is the transition of an electron from a bound state to a free state, or from one free state in the neighbourhood of an ion to another free state of greater energy. The theory hitherto used is KRAMERS’ theory of the converse process of emission by a free electron passing a positive nucleus. Since emission and absorption are intimately connected by thermodynamics, the absorption coefficient can be calculated from KRAMERS’ formulae. Unfortunately, although KRAMERS’ work is in good agreement with laboratory observations of X-rays, it gives an absorption coefficient many times smaller than that found from astronomical observations. KRAMERS used classical electromagnetism, and got over the difficulty of the quantisation of negative energies by distributing the classical emission that involved captures somewhat arbitrarily among the various stationary states. It was evidently desirable to do the same work by means of quantum theory, both for the sake of greater rigour, and in the hope of finding a larger absorption. The foundations of such a theory were laid by OPPENHEIMER,|| upon the bed-rock of SCHRODINGER’s equation, in a paper to which this one is much indebted. The matrix-elements involving positive energies present considerable difficulty, and the approximations used by OPPENHEIMER in his paper of 1927 are unsuitable for stellar applications.

scholarly journals Histidine-rich Ca2+-binding protein stimulates the transport cycle of SERCA through a conformation-dependent fuzzy complex

2020 ◽  
Author(s):  
Temitope I. Ayeotan ◽  
Line Cecilie Hansen ◽  
Thomas Boesen ◽  
Claus Olesen ◽  
Jesper V. Møller ◽  
...  
Keyword(s):  
Electrostatic Interactions ◽  
Binding Protein ◽  
Intrinsically Disordered Protein ◽  
Bound State ◽  
Disordered Proteins ◽  
Dependent Manner ◽  
Intrinsically Disordered ◽  
Intracellular Ca ◽  
P Type ◽  
Rate Limiting

AbstractThe histidine-rich Ca2+-binding protein (HRC) stimulates the sarco-endoplasmic reticulum Ca2+-ATPase (SERCA) to increase Ca2+-uptake into the lumen. HRC also binds the triadin scaffold in a Ca2+-dependent manner, and HRC tunes both the uptake and release of Ca2+ depending on the concentration in the intracellular Ca2+-stores. We investigated how HRC stimulates SERCA pumping using biochemical and biophysical assays, and show that HRC is an intrinsically disordered protein that binds directly to SERCA via electrostatic interactions. The affinity of the interaction depends on the conformation of SERCA, and HRC binds most tightly in the calcium-released E2P state. This state marks the end of the rate-limiting [Ca2]E1P to E2P transition of SERCA, and suggests that HRC stimulates SERCA by preferentially stabilizing the end point of this transition. HRC remains disordered in the bound state and thus binds in a dynamic, fuzzy complex. The binding of HRC to SERCA shows that fuzzy complexes formed by disordered proteins may be conformation-specific, and use this specificity to modulate the functional cycle of complex molecular machines such as a P-type ATPase.

scholarly journals Application of the Movable Type Free Energy Method to the Caspase-Inhibitor BindingAffinity Study

2019 ◽  
Vol 20 (19) ◽  
pp. 4850
Author(s):  
Xue ◽  
Liu ◽  
Zheng
Keyword(s):  
Free Energy ◽  
Energy Method ◽  
Structural Changes ◽  
Binding Free Energy ◽  
Bound State ◽  
Free State ◽  
Caspase Inhibitor ◽  
Caspase Inhibition ◽  
Test Sets ◽  
Inhibitor Interaction

Many studies have provided evidence suggesting that caspases not only contribute to the neurodegeneration associated with Alzheimer’s disease (AD) but also play essential roles in promoting the underlying pathology of this disease. Studies regarding the caspase inhibition draw researchers’ attention through time due to its therapeutic value in the treatment of AD. In this work, we apply the “Movable Type” (MT) free energy method, a Monte Carlo sampling method extrapolating the binding free energy by simulating the partition functions for both free-state and bound-state protein and ligand configurations, to the caspase-inhibitor binding affinity study. Two test benchmarks are introduced to examine the robustness and sensitivity of the MT method concerning the caspase inhibition complexing. The first benchmark employs a large-scale test set including more than a hundred active inhibitors binding to caspase-3. The second benchmark includes several smaller test sets studying the relative binding free energy differences for minor structural changes at the caspase-inhibitor interaction interfaces. Calculation results show that the RMS errors for all test sets are below 1.5 kcal/mol compared to the experimental binding affinity values, demonstrating good performance in simulating the caspase-inhibitor complexing. For better understanding the protein-ligand interaction mechanism, we then take a closer look at the global minimum binding modes and free-state ligand conformations to study two pairs of caspase-inhibitor complexes with (1) different caspase targets binding to the same inhibitor, and (2) different polypeptide inhibitors targeting the same caspase target. By comparing the contact maps at the binding site of different complexes, we revealed how small structural changes affect the caspase-inhibitor interaction energies. Overall, this work provides a new free energy approach for studying the caspase inhibition, with structural insight revealed for both free-state and bound-state molecular configurations.

scholarly journals Unraveling the Thermodynamics of Ultra-tight Binding of Intrinsically Disordered Proteins

2021 ◽  
Vol 8 ◽  
Author(s):  
Uroš Zavrtanik ◽  
San Hadži ◽  
Jurij Lah
Keyword(s):  
Protein Interactions ◽  
Intrinsically Disordered Proteins ◽  
Tight Binding ◽  
Bound State ◽  
Globular Proteins ◽  
Disordered Proteins ◽  
Prothymosin Α ◽  
Intrinsically Disordered ◽  
High Affinity ◽  
Entropy Optimization

Protein interactions mediated by the intrinsically disordered proteins (IDPs) are generally associated with lower affinities compared to those between globular proteins. Here, we characterize the association between the intrinsically disordered HigA2 antitoxin and its globular target HigB2 toxin from Vibrio cholerae using competition ITC experiments. We demonstrate that this interaction reaches one of the highest affinities reported for IDP-target systems (KD = 3 pM) and can be entirely attributed to a short, 20-residue-long interaction motif that folds into α-helix upon binding. We perform an experimentally based decomposition of the IDP-target association parameters into folding and binding contributions, which allows a direct comparison of the binding contribution with those from globular ultra-high affinity binders. We find that the HigA2-HigB2 interface is energy optimized to a similar extent as the interfaces of globular ultra-high affinity complexes, such as barnase-barstar. Evaluation of other ultra-high affinity IDP-target systems shows that a strategy based on entropy optimization can also achieve comparably high, picomolar affinities. Taken together, these examples show how IDP-target interactions achieve picomolar affinities either through enthalpy optimization (HigA2-HigB2), resembling the ultra-high affinity binding of globular proteins, or via bound-state fuzziness and entropy optimization (CcdA-CcdB, histone H1-prothymosin α).

scholarly journals WNK kinases sense molecular crowding and rescue cell volume via phase separation

2022 ◽  
Author(s):  
Cary R. Boyd-Shiwarski ◽  
Daniel J. Shiwarski ◽  
Shawn E. Griffiths ◽  
Rebecca T. Beacham ◽  
Logan Norrell ◽  
...  
Keyword(s):  
Phase Separation ◽  
Cell Volume ◽  
Molecular Crowding ◽  
Intrinsically Disordered ◽  
Physiological Constraints ◽  
C Terminus ◽  
Dependent Signal ◽  
Wnk Kinases ◽  
A Cell

When challenged by hypertonicity, dehydrated cells must defend their volume to survive. This process requires the phosphorylation-dependent regulation of SLC12 cation chloride transporters by WNK kinases, but how these kinases are activated by cell shrinkage remains unknown. Within seconds of cell exposure to hypertonicity, WNK1 concentrates into membraneless droplets, initiating a phosphorylation-dependent signal that drives net ion influx via the SLC12 cotransporters to rescue volume. The formation of WNK1 condensates is driven by its intrinsically disordered C-terminus, whose evolutionarily conserved signatures are necessary for efficient phase separation and volume recovery. This disorder-encoded phase behavior occurs within physiological constraints and is activated in vivo by molecular crowding rather than changes in cell size. This allows WNK1 to bypass a strengthened ionic milieu that favors kinase inactivity and reclaim cell volume through condensate-mediated signal amplification. Thus, WNK kinases are physiological crowding sensors that phase separate to coordinate a cell volume rescue response.

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