scholarly journals Divining Deamidation and Isomerization in Therapeutic Proteins: Effect of Neighboring Residue

2021 ◽  
Author(s):  
Flaviyan Jerome Irudayanathan ◽  
Jonathan Zarzar ◽  
Jasper Lin ◽  
Saeed Izadi
Keyword(s):  
Aspartic Acid ◽  
Proton Affinity ◽  
Solvent Accessibility ◽  
Clinical Stage ◽  
Therapeutic Proteins ◽  
Mechanistic Explanation ◽  
Md Simulations ◽  
Classification Model ◽  
Semi Empirical ◽  
Molecular Indicators

Deamidation of asparagine (ASN) and isomerization of aspartic acid (ASP) residues are among the most commonly observed spontaneous post-translational modifications (PTMs) in proteins. Understanding and predicting a protein sequence's propensity for such PTMs can help expedite protein therapeutic discovery and development. In this study, we utilized proton-affinity calculations with semi-empirical quantum mechanics (QM) and microsecond long equilibrium molecular dynamics (MD) simulations to investigate mechanistic roles of structure and chemical environment in dictating spontaneous degradation of asparagine and aspartic acid residues in 131 clinical-stage therapeutic antibodies. Backbone secondary structure, side-chain rotamer conformation and solvent accessibility were found as three key molecular indicators of ASP isomerization and ASN deamidation. Comparative analysis of backbone dihedral angles along with N-H proton affinity calculations provides a mechanistic explanation for the strong influence of the identity of the n+1 residue on the rate of ASP/ASN degradation. With these findings, we propose a minimalistic physics-based classification model that can be leveraged to predict deamidation and isomerization propensity of therapeutic proteins.

scholarly journals In silico Evaluation of Cucurbit[6]uril as a Potential Detector for Cocaine and Its Adulterants Lidocaine, Caffeine, and Procaine

2021 ◽  
Author(s):  
Caio Rodrigues ◽  
Jorge Hernández-González ◽  
Natalia Pedrina ◽  
Vitor Leite ◽  
Aline Bruni
Keyword(s):  
In Silico ◽  
Density Functional ◽  
Illicit Drugs ◽  
Binding Free Energy ◽  
Md Simulations ◽  
Basis Set ◽  
Phase Condition ◽  
Cocaine Detection ◽  
Semi Empirical ◽  
And Control

Illicit drugs and their trafficking require worldwide efforts in investigation, detection, and control. Colorimetric tests are often applied to identify drugs. Cocaine has some well-known adulterants that can provide a false positive response. Cucurbit[6]uril (CB[6]) has been suggested as a potential detector for cocaine and other illicit drugs. This work uses in silico methods to evaluate the use of CB[6] to detect cocaine and these interfering substances. More specifically, this work analyzes different possibilities of CB[6] complexation with cocaine, lidocaine, caffeine, and procaine and compares the results achieved for cocaine and its adulterants. Different methodologies were employed: quantum chemistry was investigated through DFT B3LYP/TZVP (density functional theory-Becke, three-parameter, Lee-Yang-Parr with triple zeta valence plus polarization basis set) and the semi-empirical methods Austin model 1 (AM1), parametric methods 3, 6, and 7 (PM3, PM6, PM7), and Recife model 1 (RM1). We used these methodologies intending to compare the reasonability and reproducibility of the results in the gas phase condition. Solvent influence was studied by molecular dynamics (MD) simulations. Results showed that CB[6] does not bind to these substances, as judged from the positive values of binding free energy obtained with all methods. DFT and MD were the most reliable methods whereas semiempirical ones were not reproductible in describing these systems. Results also showed that interactions are not specific, so CB[6] does not provide a good response for cocaine detection.

scholarly journals 3D Interaction Homology: Computational Titration of Aspartic Acid, Glutamic Acid and Histidine Can Create pH-Tunable Hydropathic Environment Maps

2021 ◽  
Vol 8 ◽  
Author(s):  
Noah B. Herrington ◽  
Glen E. Kellogg
Keyword(s):  
Glutamic Acid ◽  
Aspartic Acid ◽  
Protein Design ◽  
Solvent Accessibility ◽  
Scoring Function ◽  
Relative Solvent Accessibility ◽  
Solvent Exposure ◽  
3D Interaction ◽  
Ionizable Residues ◽  
Environment Maps

Aspartic acid, glutamic acid and histidine are ionizable residues occupying various protein environments and perform many different functions in structures. Their roles are tied to their acid/base equilibria, solvent exposure, and backbone conformations. We propose that the number of unique environments for ASP, GLU and HIS is quite limited. We generated maps of these residue's environments using a hydropathic scoring function to record the type and magnitude of interactions for each residue in a 2703-protein structural dataset. These maps are backbone-dependent and suggest the existence of new structural motifs for each residue type. Additionally, we developed an algorithm for tuning these maps to any pH, a potentially useful element for protein design and structure building. Here, we elucidate the complex interplay between secondary structure, relative solvent accessibility, and residue ionization states: the degree of protonation for ionizable residues increases with solvent accessibility, which in turn is notably dependent on backbone structure.

scholarly journals Quantum Chemical Methods for Modeling Covalent Modification of Biological Thiols

2019 ◽  
Author(s):  
Ernest Awoonor-Williams ◽  
William Isley ◽  
Stephen Dale ◽  
Erin Johnson ◽  
Haibo Yu ◽  
...  
Keyword(s):  
Density Functional ◽  
Functional Performance ◽  
Low Cost ◽  
Md Simulations ◽  
Potential Of Mean Force ◽  
Covalent Modification ◽  
Dft Methods ◽  
Aqueous Solvent ◽  
Exact Exchange ◽  
Semi Empirical

Targeted covalent inhibitor drugs require computational methods that go beyond simple molecular-mechanical force fields in order to model the chemical reactions that occur when they bind to their targets. Here, several semi-empirical and density-functional theory (DFT) methods are assessed for their ability to describe the potential energy surface and reaction energies of the covalent modification of a thiol by an electrophile. Functionals such as PBE and B3LYP fail to predict a stable enolate intermediate. This is largely due to delocalization error, which spuriously stabilizes the pre-reaction complex, in which excess electron density is transferred from the thiolate to the electrophile. Functionals with a high-exact exchange component, range-separated DFT functionals, and variationally-optimized exact exchange (i.e., the LC-B05minV functional) correct this issue to various degrees. The large gradient behaviour of the exchange enhancement factor is also found to significantly affect the results, leading to the improved performance of PBE0. While ωB97X-D and M06-2X were easonably accurate, no method provided quantitative accuracy for all three electrophiles, making this a very strenuous test of functional performance. Additionally, one drawback of M06-2X was that MD simulations using this functional were only stable if a fine integration grid was used. The low-cost semi-empirical methods, PM3, AM1, and PM7, provide a qualitatively correct description of the reaction mechanism, although the energetics are not quantitatively reliable. As a proof of concept, the potential of mean force for the addition of methylthiolate to MVK was calculated using QM/MM MD in an explicit polarizable aqueous solvent.<br>

scholarly journals Theoretical Inversion of Amino Acids (Alanine and Aspartic Acid) by Semi-Empirical Methods

2013 ◽  
Vol 12 ◽  
pp. 37-44
Author(s):  
Musa E. Mohamed
Keyword(s):  
Activation Energy ◽  
Amino Acids ◽  
Transition State ◽  
Gas Phase ◽  
Aspartic Acid ◽  
Aqueous Phase ◽  
Free Amino ◽  
Membered Ring ◽  
Empirical Methods ◽  
Semi Empirical

The inversion reaction coordinate of free amino acids (alanine, aspartic acid) have been computationally calculated by semi-empirical methods AM1. A transition state for free alanine and aspartic acid were obtained as a three membered ring in which the α-C-H and α-C-CH3 are slightly elongated, 1.2 and 2.17 Å respectively in the alanine transition state. The activation energy of alanine is 77.52 kcal/mol in the gas phase and 76.66 kcal/mol in aqueous phase, and for aspartic acid is 54.87 kcal/mol in the gas phase and 50.86 kcal/mol in aqueous phase.

scholarly journals Radical Stabilization Energies for Enzyme Engineering – Tackling the Substrate Scope of the Radical Enzyme QueE

2018 ◽  
Author(s):  
Christian Suess ◽  
Floriane Martins ◽  
Anna Croft ◽  
Christof Jaeger
Keyword(s):  
Screening Method ◽  
Cost Effective ◽  
Md Simulations ◽  
Enzyme Engineering ◽  
Radical Intermediates ◽  
Stabilization Energies ◽  
Effective Assessment ◽  
Semi Empirical ◽  
The Impact ◽  
Radical Enzymes

Experimental assessment of the reaction mechanisms and profiles of radical enzymes can be severely challenging due to the reactive nature of the intermediates, and sensitivity of cofactors such as iron sulfur clusters. Here we present an enzyme-directed computational methodology for the assessment of thermodynamic reaction profiles and screening for radical stabilization energies (RSEs) for the assessment of catalytic turnovers in radical enzymes. We have applied this new screening method to the radical SAM enzyme CPH4 synthase (QueE), following a detailed molecular dynamics (MD) analysis that clarifies the role of both specific enzyme residues and bound Mg2+, Ca2+ or Na+. The MD simulations provided the basis for a statistical approach to sample different conformational outcomes. RSE calculation at the M06-2X/6-31+G* level of theory provided the most computationally cost-effective assessment of enzyme-based energies, facilitated by an initial triage using semi-empirical methods. The impact of intermolecular interactions on RSE was clearly established and application to the assessment of potential alternative substrates (focusing on radical clock type rearrangements) proposes a selection of carbon-substituted analogues that would react to afford cyclopropylcarbinyl radical intermediates, as candidates for catalytic turnover by QueE.

scholarly journals Quantum Chemical Methods for Modeling Covalent Modification of Biological Thiols

2019 ◽  
Author(s):  
Ernest Awoonor-Williams ◽  
William Isley ◽  
Stephen Dale ◽  
Erin Johnson ◽  
Haibo Yu ◽  
...  
Keyword(s):  
Density Functional ◽  
Functional Performance ◽  
Low Cost ◽  
Md Simulations ◽  
Potential Of Mean Force ◽  
Covalent Modification ◽  
Dft Methods ◽  
Aqueous Solvent ◽  
Exact Exchange ◽  
Semi Empirical

Targeted covalent inhibitor drugs require computational methods that go beyond simple molecular-mechanical force fields in order to model the chemical reactions that occur when they bind to their targets. Here, several semi-empirical and density-functional theory (DFT) methods are assessed for their ability to describe the potential energy surface and reaction energies of the covalent modification of a thiol by an electrophile. Functionals such as PBE and B3LYP fail to predict a stable enolate intermediate. This is largely due to delocalization error, which spuriously stabilizes the pre-reaction complex, in which excess electron density is transferred from the thiolate to the electrophile. Functionals with a high-exact exchange component, range-separated DFT functionals, and variationally-optimized exact exchange (i.e., the LC-B05minV functional) correct this issue to various degrees. The large gradient behaviour of the exchange enhancement factor is also found to significantly affect the results, leading to the improved performance of PBE0. While ωB97X-D and M06-2X were easonably accurate, no method provided quantitative accuracy for all three electrophiles, making this a very strenuous test of functional performance. Additionally, one drawback of M06-2X was that MD simulations using this functional were only stable if a fine integration grid was used. The low-cost semi-empirical methods, PM3, AM1, and PM7, provide a qualitatively correct description of the reaction mechanism, although the energetics are not quantitatively reliable. As a proof of concept, the potential of mean force for the addition of methylthiolate to MVK was calculated using QM/MM MD in an explicit polarizable aqueous solvent.<br>

scholarly journals Interplay of folded domains and the disordered low-complexity domain in mediating hnRNPA1 phase separation

2020 ◽  
Author(s):  
Erik W. Martin ◽  
F. Emil Thomasen ◽  
Nicole M. Milkovic ◽  
Matthew J. Cuneo ◽  
Christy R. Grace ◽  
...  
Keyword(s):  
Phase Separation ◽  
Phase Behavior ◽  
Rna Binding ◽  
Mechanistic Explanation ◽  
Md Simulations ◽  
Low Complexity ◽  
Coarse Grained ◽  
Intrinsically Disordered ◽  
X Ray Scattering

AbstractLiquid-liquid phase separation underlies the membrane-less compartmentalization of cells. Intrinsically disordered low-complexity domains (LCDs) often mediate phase separation, but how their phase behavior is modulated by folded domains is incompletely understood. Here, we interrogate the interplay between folded and disordered domains of the RNA-binding protein hnRNPA1. The LCD of hnRNPA1 is sufficient for mediating phase separation in vitro. However, we show that the folded RRM domains and a folded solubility-tag modify the phase behavior, even in the absence of RNA. Notably, the presence of the folded domains reverses the salt dependence of the driving force for phase separation relative to the LCD alone. Small-angle X-ray scattering experiments and coarse-grained MD simulations show that the LCD interacts transiently with the RRMs and/or the solubility-tag in a salt-sensitive manner, providing a mechanistic explanation for the observed salt-dependent phase separation. These data point to two effects from the folded domains: (1) electrostatically mediated interactions that compact hnRNPA1 and contribute to phase separation, and (2) increased solubility at higher ionic strengths mediated by the folded domains. The interplay between disordered and folded domains can modify the dependence of phase behavior on solution conditions and can obscure signatures of physicochemical interactions underlying phase separation.Graphical abstracthnRNPA1 phase separation is highly salt sensitive.Phase separation of the low-complexity domain (LCD) of hnRNPA1 increases with NaCl. In contrast, phase separation of full-length hnRNPA1 is saltsensitive. At low NaCl concentrations, electrostatic RRM-LCD interactions occur and can contribute positively to phase separation, but they are screened at high NaCl concentrations. The folded domains solubilize hnRNPA1 under these conditions and prevent phase separation.

scholarly journals Radical Stabilization Energies for Enzyme Engineering – Tackling the Substrate Scope of the Radical Enzyme QueE

2019 ◽  
Author(s):  
Christian Suess ◽  
Floriane Martins ◽  
Anna Croft ◽  
Christof Jaeger
Keyword(s):  
Screening Method ◽  
Cost Effective ◽  
Md Simulations ◽  
Enzyme Engineering ◽  
Radical Intermediates ◽  
Stabilization Energies ◽  
Effective Assessment ◽  
Semi Empirical ◽  
The Impact ◽  
Radical Enzymes

Experimental assessment of the reaction mechanisms and profiles of radical enzymes can be severely challenging due to the reactive nature of the intermediates, and sensitivity of cofactors such as iron sulfur clusters. Here we present an enzyme-directed computational methodology for the assessment of thermodynamic reaction profiles and screening for radical stabilization energies (RSEs) for the assessment of catalytic turnovers in radical enzymes. We have applied this new screening method to the radical SAM enzyme CPH4 synthase (QueE), following a detailed molecular dynamics (MD) analysis that clarifies the role of both specific enzyme residues and bound Mg2+, Ca2+ or Na+. The MD simulations provided the basis for a statistical approach to sample different conformational outcomes. RSE calculation at the M06-2X/6-31+G* level of theory provided the most computationally cost-effective assessment of enzyme-based energies, facilitated by an initial triage using semi-empirical methods. The impact of intermolecular interactions on RSE was clearly established and application to the assessment of potential alternative substrates (focusing on radical clock type rearrangements) proposes a selection of carbon-substituted analogues that would react to afford cyclopropylcarbinyl radical intermediates, as candidates for catalytic turnover by QueE.

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