scholarly journals Nature of Allosteric Inhibition in Glutamate Racemase: Discovery and Characterization of a Cryptic Inhibitory Pocket Using Atomistic MD Simulations and pKaCalculations

2011 ◽  
Vol 115 (13) ◽  
pp. 3416-3424 ◽  
Author(s):  
Katie L. Whalen ◽  
Kenneth B. Tussey ◽  
Steven R. Blanke ◽  
M. Ashley Spies
Keyword(s):  
Md Simulations ◽  
Allosteric Inhibition ◽  
Glutamate Racemase

Prediction and characterization of the stability enhancing effect of the Cherry-Tag™ in highly concentrated protein solutions by complex rheological measurements and MD simulations

2017 ◽  
Vol 531 (1) ◽  
pp. 360-371 ◽  
Author(s):  
Pascal Baumann ◽  
Marie-Therese Schermeyer ◽  
Hannah Burghardt ◽  
Cathrin Dürr ◽  
Jonas Gärtner ◽  
...  
Keyword(s):  
Md Simulations ◽  
Protein Solutions ◽  
Enhancing Effect ◽  
Rheological Measurements ◽  
The Stability

scholarly journals Biophysical characterization of the SARS-CoV-2 E protein

2021 ◽  
Author(s):  
Aujan Mehregan ◽  
Sergio Perez-Conesa ◽  
Yuxuan Zhuang ◽  
Ahmad Elbahnsi ◽  
Diletta Pasini ◽  
...  
Keyword(s):  
Recombinant Expression ◽  
Structural Data ◽  
Md Simulations ◽  
Full Length ◽  
Coarse Grained ◽  
Biophysical Characterization ◽  
Viral Budding ◽  
Nmr Structures ◽  
E Protein

SARS-CoV-2 is the virus responsible for the COVID-19 pandemic which continues to wreak havoc across the world, over a year and a half after its effects were first reported in the general media. Current fundamental research efforts largely focus on the SARS-CoV-2 Spike protein. Since successful antiviral therapies are likely to target multiple viral components, there is considerable interest in understanding the biophysical role of its other proteins, in particular structural membrane proteins. Here, we have focused our efforts on the characterization of the full-length E protein from SARS-CoV-2, combining experimental and computational approaches. Recombinant expression of the full-length E protein from SARS-CoV-2 reveals that this membrane protein is capable of independent multimerization, possibly as a tetrameric or smaller species. Fluorescence microscopy shows that the protein localizes intracellularly, and coarse-grained MD simulations indicate it causes bending of the surrounding lipid bilayer, corroborating a potential role for the E protein in viral budding. Although we did not find robust electrophysiological evidence of ion-channel activity, cells transfected with the E protein exhibited reduced intracellular Ca2+, which may further promote viral replication. However, our atomistic MD simulations revealed that previous NMR structures are relatively unstable, and result in models incapable of ion conduction. Our study highlights the importance of using high-resolution structural data obtained from a full-length protein to gain detailed molecular insights, and eventually permitting virtual drug screening.

scholarly journals Engineering P450 TamI as an Iterative Biocatalyst for Selective Late-Stage C-H Functionalization and Epoxidation of Tirandamycin Antibiotics

2021 ◽  
Author(s):  
Rosa V. Espinoza ◽  
Kersti Caddell Haatveit ◽  
S. Wald Grossman ◽  
Jin Yi Tan ◽  
Caylie A. McGlade ◽  
...  
Keyword(s):  
Natural Product ◽  
Late Stage ◽  
Md Simulations ◽  
Cascade Reactions ◽  
Direct Access ◽  
Tetramic Acid ◽  
Reaction Sequence ◽  
Substrate Reactivity ◽  
Continuous Oxidation

<div> <div> <div> <p>Iterative P450 enzymes are powerful biocatalysts for selective late-stage C-H oxidation of complex natural product scaffolds. These enzymes represent new tools for selectivity and cascade reactions, facilitating direct access to core structure diversification. Recently, we reported the structure of the multifunctional bacterial P450 TamI and elucidated the molecular basis of its substrate binding and strict reaction sequence at distinct carbon atoms of the substrate. Here, we report the design and characterization of a toolbox of TamI biocatalysts, generated by mutations at Leu101, Leu244 and/or Leu295, that alter the native selectivity, step sequence and number of reactions catalyzed, including the engineering of a variant capable of catalyzing a four-step oxidative cascade without the assistance of the flavoprotein and oxidative partner TamL. The tuned enzymes override inherent substrate reactivity enabling catalyst- controlled C-H functionalization and alkene epoxidation of the tetramic acid-containing natural product tirandamycin. Five new, bioactive tirandamycin derivatives (6-10) were generated through TamI-mediated enzymatic synthesis. Quantum mechanics calculations and MD simulations provide important insights on the basis of altered selectivity and underlying biocatalytic mechanisms for enhanced continuous oxidation of the iterative P450 TamI. </p> </div> </div> </div>

scholarly journals Atomic-level characterization of protein–protein association

2019 ◽  
Vol 116 (10) ◽  
pp. 4244-4249 ◽  
Author(s):  
Albert C. Pan ◽  
Daniel Jacobson ◽  
Konstantin Yatsenko ◽  
Duluxan Sritharan ◽  
Thomas M. Weinreich ◽  
...  
Keyword(s):  
Structural Information ◽  
Protein Complexes ◽  
Md Simulations ◽  
Atomic Level ◽  
Conformational Ensemble ◽  
Native State ◽  
Protein Interfaces ◽  
Study Association ◽  
Functionally Diverse

Despite the biological importance of protein–protein complexes, determining their structures and association mechanisms remains an outstanding challenge. Here, we report the results of atomic-level simulations in which we observed five protein–protein pairs repeatedly associate to, and dissociate from, their experimentally determined native complexes using a molecular dynamics (MD)–based sampling approach that does not make use of any prior structural information about the complexes. To study association mechanisms, we performed additional, conventional MD simulations, in which we observed numerous spontaneous association events. A shared feature of native association for these five structurally and functionally diverse protein systems was that if the proteins made contact far from the native interface, the native state was reached by dissociation and eventual reassociation near the native interface, rather than by extensive interfacial exploration while the proteins remained in contact. At the transition state (the conformational ensemble from which association to the native complex and dissociation are equally likely), the protein–protein interfaces were still highly hydrated, and no more than 20% of native contacts had formed.

scholarly journals Monitoring Unfolding of Titin I27 single- and bi-Domain with High-Pressure NMR Spectroscopy

2018 ◽  
Author(s):  
I. Herrada ◽  
P. Barthe ◽  
M. Van Heusden ◽  
K DeGuillen ◽  
L Mammri ◽  
...  
Keyword(s):  
High Pressure ◽  
Force Spectroscopy ◽  
Md Simulations ◽  
Specific Information ◽  
Folding Energy ◽  
3D Scaffold ◽  
Folding Process ◽  
Energetic Characterization ◽  
Volume Kinetics

ABSTRACTA complete description of the pathways and mechanisms of protein folding requires a detailed structural and energetic characterization of the folding energy landscape. Simulations, when corroborated by experimental data yielding global information on the folding process, can provide this level of insight. Molecular Dynamics (MD) has been associated often to force spectroscopy experiments to decipher the unfolding mechanism of titin Ig-like single- or multi-domain, the giant multi-modular protein from sarcomere, yielding information on the sequential events during titin unfolding under stretching. Here, we used high-pressure NMR to monitor the unfolding of titin I27 Ig-like single-domain and tandem. Since this method brings residue-specific information on the folding process, it can provide quasi-atomic details on this process, without the help of MD simulations. Globally, the results of our high-pressure analysis are in agreement with previous results obtained by the association of experimental measurements and MD simulation and/or protein engineering, although the intermediate folding state caused by the early detachment of the AB ß-sheet, often reported in previous works based on MD or force spectroscopy, cannot be detected. On the other hand, the A’G parallel ß-sheet of the ß-sandwich has been confirmed as the Achilles heel of the 3D scaffold: its disruption yields complete unfolding, with very similar characteristics (free energy, unfolding volume, kinetics constant rates) for the two constructs.

scholarly journals Expression, purification and characterization of anti-FGF7 domain antibody identified using phage display technique

2021 ◽  
Author(s):  
Aliakbar Alizadeh ◽  
Mona Roshani ◽  
Omid Jamshidi Kandjani ◽  
Milad Soltani-Saif ◽  
Siavoush Dastmalchi
Keyword(s):  
Phage Display ◽  
Md Simulations ◽  
Docking Study ◽  
Single Domain Antibody ◽  
Cancer Cell Growth ◽  
Phage Display Technique ◽  
Domain Antibody ◽  
Sds Page ◽  
Display Technique

Background: Fibroblast growth factors (FGFs) are involved in angiogenesis, wound healing and embryonic development. However, one of the causes of cancer cell growth in fibroblast-dependent cancers is FGF7 secreted by fibroblasts. Therefore, antibodies against FGF7 can be used for treatment of these types of cancers. Methods: In previous studies, a phage displaying single domain antibody, D53, against human FGF7 has been identified using the phage display technique. In the present study, D53 was produced and purified in its isolated form. ELISA experiment was performed to evaluate the binding of D53 to FGF7. The mode of interaction of D53-FGF7 was explored using docking study and molecular dynamics (MD) simulations. Results: The expression and purification processes were verified using western blotting and SDS-PAGE analyses. ELISA experiment showed that D53 is able to recognize and bind FGF7. Docking study and MD simulations indicated that compared to dummy VH, D53 has more affinity towards FGF7. Conclusion: The findings in the current study can be useful for generation and development of FGF7 inhibitors with potential use in fibroblast-dependent cancers.

scholarly journals From Crystallographic Data to Solution Structure of Photoreceptors: The Case of AppA BLUF Domain

2021 ◽  
Author(s):  
Shaima Hashem ◽  
Veronica Macaluso ◽  
Michele Nottoli ◽  
Filippo Lipparini ◽  
Lorenzo Cupellini ◽  
...  
Keyword(s):  
Solution Structure ◽  
Molecular Mechanisms ◽  
Md Simulations ◽  
Hydrogen Bond Network ◽  
Light Activation ◽  
Activated State ◽  
Bond Network ◽  
Bluf Domain ◽  
First Time

<div>Photoreceptor proteins bind a chromophore, which, upon light absorption, modifies its geometry or its interactions with the protein, finally inducing the structural change needed to switch the protein from an inactive to an active or signaling state. In the Blue Light-Using Flavin (BLUF) family of photoreceptors, the chromophore is a flavin and the changes have been connected with a rearrangement of the hydrogen bond network around it on the basis of spectroscopic changes measured for the dark-to-light conversion. However, the exact conformational change triggered by the photoexcitation is still elusive mainly because a clear consensus on the identity not only of the light activated state but also of the dark one has not been achieved. Here, for the first time, we present an integrated investigation that combines microsecond MD simulations starting from the two conflicting crystal structures available for the AppA BLUF domain with calculations of NMR, IR and UV-Vis spectra using a polarizable QM/MM approach. Thanks to such a combined analysis of the three different spectroscopic responses, a robust characterization of the structure of the dark state in solution is given together with the uncovering of important flaws of the most popular molecular mechanisms present in the literature for the dark-to-light activation. </div>

Stability and characterization of the structure II binary clathrate hydrate of the refrigerant trans-1,3,3,3-tetrafluoropropene + methane

2019 ◽  
Vol 43 (33) ◽  
pp. 13068-13074 ◽  
Author(s):  
Kotaro Nemoto ◽  
Takumi Ikeda ◽  
Hiroyuki Mori ◽  
Saman Alavi ◽  
Satoshi Takeya ◽  
...  
Keyword(s):  
Phase Equilibrium ◽  
Md Simulations ◽  
Clathrate Hydrate

A new clathrate hydrate formed with trans-1,3,3,3-tetrafluoropropene and methane was characterized by phase equilibrium and PXRD measurements and MD simulations.

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