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1.
Phys Chem Chem Phys ; 21(36): 19795-19804, 2019 Sep 18.
Artigo em Inglês | MEDLINE | ID: mdl-31482888

RESUMO

Allostery plays important roles in the regulation of many biological processes, such as signal transduction and transcriptional regulation. Although great advances have been achieved in understanding the allosteric mechanism through experimental and theoretical investigations, the details of the allosteric process are still not clear. Here, using the N-terminal domain of calmodulin (nCaM) as the model protein, we reported the atomic level characterization of the allosteric process induced by Ca2+ binding through extensive and unbiased molecular dynamics simulations. In two trajectories, it was found that Ca2+ first binds to EF-hand 2 and then induces the conformational transformation of nCaM from the Apo to Holo state assisted by second Ca2+ binding to EF-hand 1 completely. The binding order was consistent with a recent experimental result. The simulations also indicated that the two EF-hands changed conformations synergistically and the EF-hand 2 showed an earlier and more gradual conformational transition. Meanwhile, the allosteric process of nCaM triggered by Ca2+ binding might be completed within hundreds of nanoseconds in a two-state-like manner. This was validated by biased simulations, in which the Ca2+ ions were restrained near the binding sites. This work provides the molecular details of the conformational transition of nCaM triggered by Ca2+ binding.


Assuntos
Cálcio/química , Calmodulina/química , Íons/química , Simulação de Acoplamento Molecular , Domínios Proteicos , Ligação Proteica , Conformação Proteica
2.
J Chem Phys ; 151(8): 085101, 2019 Aug 28.
Artigo em Inglês | MEDLINE | ID: mdl-31470695

RESUMO

Periodic molecular dynamics simulations of proteins may suffer from image interactions. Similarly, the hydrophobic effect required to keep a protein folded may not be enforced by small simulation cells. Accordingly, errors may arise both from the water concentration per se and the image interactions. Intrinsically disordered proteins are particularly sensitive, providing a worst-case estimate of the errors. Following this reasoning, we studied Aß40 (Aß), a disordered peptide central to Alzheimer's disease, by 100 different simulations with variable cell size from very large (20 Å) to very small (3 Å). Even for this very disordered peptide, most properties are not cell-size dependent, justifying the common use of modest-sized (10 Å) cells for simulating proteins. The radius of gyration, secondary structure, intrapeptide, and peptide-water hydrogen bonds are similar relative to standard deviations at any cell size. However, hydrophobic surface area increases significantly in small cells (confidence 95%, two-tailed t-test), as does the standard deviation in exposure and backbone conformations (>40% and >27%). Similar results were obtained for the force fields OPLS3e, Ambersb99-ILDN, and Charmm22*. The similar prevalence of structures and α-ß transitions in long and short simulations indicate small diffusion barriers, which we suggest is a defining hallmark of intrinsically disordered proteins. Whereas hydrophilic exposure dominates in large cells, hydrophobic exposure dominates in small cells, suggesting a weakening of the hydrophobic effect by image interactions and the few water layers available to keep the protein compact, with a critical limit of 2-3 water layers required to enforce the hydrophobic effect.


Assuntos
Peptídeos beta-Amiloides/química , Tamanho Celular , Proteínas Intrinsicamente Desordenadas/química , Simulação de Dinâmica Molecular , Humanos , Interações Hidrofóbicas e Hidrofílicas , Conformação Proteica
3.
Hum Genet ; 138(10): 1183-1200, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31471722

RESUMO

The glutamate pyruvate transaminase 2 (GPT2) gene produces a nuclear-encoded mitochondrial enzyme that catalyzes the reversible transfer of an amino group from glutamate to pyruvate, generating alanine and alpha-ketoglutarate. Recessive mutations in GPT2 have been recently identified in a new syndrome involving intellectual and developmental disability (IDD), postnatal microcephaly, and spastic paraplegia. We have identified additional families with recessive GPT2 mutations and expanded the phenotype to include small stature. GPT2 loss-of-function mutations were identified in four families, nine patients total, including: a homozygous mutation in one child [c.775T>C (p.C259R)]; compound heterozygous mutations in two siblings [c.812A>C (p.N271T)/c.1432_1433delGT (p.V478Rfs*73)]; a novel homozygous, putative splicing mutation [c.1035C>T (p.G345=)]; and finally, a recurrent mutation, previously identified in a distinct family [c.1210C>T (p.R404*)]. All patients were diagnosed with IDD. A majority of patients had remarkably small stature throughout development, many < 1st percentile for height and weight. Given the potential biological function of GPT2 in cellular growth, this phenotype is strongly suggestive of a newly identified clinical susceptibility. Further, homozygous GPT2 mutations manifested in at least 2 of 176 families with IDD (approximately 1.1%) in a Pakistani cohort, thereby representing a relatively common cause of recessive IDD in this population, with recurrence of the p.R404* mutation in this population. Based on variants in the ExAC database, we estimated that approximately 1 in 248 individuals are carriers of moderately or severely deleterious variants in GPT2.


Assuntos
Deficiências do Desenvolvimento/diagnóstico , Deficiências do Desenvolvimento/genética , Genes Recessivos , Predisposição Genética para Doença , Mutação , Fenótipo , Transaminases/genética , Adolescente , Alelos , Substituição de Aminoácidos , Deficiências do Desenvolvimento/metabolismo , Ativação Enzimática , Éxons , Feminino , Frequência do Gene , Estudos de Associação Genética , Genética Populacional , Genótipo , Humanos , Deficiência Intelectual/diagnóstico , Deficiência Intelectual/genética , Imagem por Ressonância Magnética , Masculino , Mitocôndrias/genética , Mitocôndrias/metabolismo , Modelos Moleculares , Linhagem , Conformação Proteica , Sítios de Splice de RNA , Análise de Sequência de DNA , Relação Estrutura-Atividade , Transaminases/química , Transaminases/metabolismo
4.
Gene ; 720: 144082, 2019 Dec 15.
Artigo em Inglês | MEDLINE | ID: mdl-31476406

RESUMO

The enzyme ß-Ketoacyl ACP synthase I (KasA) is a potent drug target in mycolic acid pathway of Mycobacterium tuberculosis (Mtb). In the present study, we investigated the structural dynamics of wild-type (WT) and mutants KasA (D66N, G269S, G312S, and F413L) in both monomer and dimer form to provide insight into protein structural stability. To gain better understanding of structural flexibility of KasA, combined molecular dynamics and essential dynamics were employed to analyze the conformational changes induced by non-active site mutations. The results confirm that non-active site mutations lower the structural stability in dimer KasA as compared to WT. The protein network topology and close residue interactions of WT and mutant residues of KasA have been predicted through residue interaction network analysis (RIN). Non-active site mutations distort RIN architecture and subsequently affect the drug binding landscape. T-pad associated with mode vector analysis comprehensively pronounces the structural impact caused by non-active site mutations. It also identified the critical fluctuating residues present in the gate segment (GS) region (115-147). The non-active site mutations altered the structural stability of the mutant protein structures, and these mutations may be a cause for the resistance mechanism of KasA against anti-tuberculosis drugs. Further, it is observed that dimer mutant KasA proteins display much more structural flexibility than WT at the ligand binding site which is evident from the binding site analysis and hydrogen bond interaction patterns. This study provides a better understanding of the structural dynamic behaviour of KasA mutants, thereby facilitating the need to find a novel and potent inhibitor against Mtb.


Assuntos
3-Oxoacil-(Proteína de Transporte de Acila) Sintase/química , Proteínas de Bactérias/química , Isoenzimas/química , Proteínas Mutantes/química , Mutação , Mycobacterium tuberculosis/enzimologia , Tuberculose/microbiologia , 3-Oxoacil-(Proteína de Transporte de Acila) Sintase/genética , Proteínas de Bactérias/genética , Isoenzimas/genética , Simulação de Dinâmica Molecular , Proteínas Mutantes/genética , Conformação Proteica , Tuberculose/genética , Tuberculose/metabolismo
5.
Phys Chem Chem Phys ; 21(32): 17893-17900, 2019 Aug 15.
Artigo em Inglês | MEDLINE | ID: mdl-31380529

RESUMO

The dispersion interaction was reported to play a critical role in the stabilization of model dipeptide Z-Arg-OH, even greater than the conventional hydrogen bond (HB), which is opposite to the traditional opinion. Here the conformation of Z-Arg-OH has been systematically searched by the effective fragment based step-by-step strategy. All the newly-found low-energy conformers determined at the advanced DSD-PBEP86-D3(BJ)/aug-cc-pVTZ level are clearly in the stretched form with strong conventional HBs, rather than the reported folded structures with emphasis on the dispersion interactions. The simulated IR spectra of the stretched conformers fit better than those of the folded ones compared with the previous experimental observations. Near-edge X-ray absorption fine-structure (NEXAFS) spectra and X-ray photoelectron spectra (XPS) at C, N and O K-edges have also been simulated to unambiguously identify different isomers. This work thus provides valuable insight into the competitions between the conventional HB and the dispersion interactions and demonstrates that the conventional hydrogen bonding is still more important for such small peptides.


Assuntos
Arginina/análogos & derivados , Arginina/química , Dipeptídeos/química , Modelos Moleculares , Ligações de Hidrogênio , Isomerismo , Fenômenos Físicos , Conformação Proteica , Estabilidade Proteica , Solventes/química , Termodinâmica
6.
J Chem Theory Comput ; 15(9): 5103-5115, 2019 Sep 10.
Artigo em Inglês | MEDLINE | ID: mdl-31402649

RESUMO

Intrinsically disordered proteins (IDPs) play key roles in biology and disease, rationalizing the wide interest in deriving accurate solution ensembles of IDPs. Molecular dynamics (MD) simulations of IDPs often suffer from force-field inaccuracies, suggesting that simulations must be complemented by experimental data to obtain physically correct ensembles. We present a method for integrating small-angle X-ray scattering (SAXS) data on-the-fly into MD simulations of disordered systems, with the aim to bias the conformational sampling toward agreement with ensemble-averaged SAXS data. By coupling a set of parallel replicas to the data and following the principle of maximum entropy, this method applies only a minimal bias. Using the RS peptide as a test case, we analyze the influence of (i) the number of parallel replicas, (ii) the scaling of the force constant for the SAXS-derived biasing energy with the number of parallel replicas, and (iii) the force field. The refined ensembles are cross-validated against experimental 3JHN-Hα couplings and further compared in terms of Cα distance maps and secondary structure content. Remarkably, we find that the applied force field only has a small influence on the SAXS-refined ensemble, suggesting that incorporating SAXS data into MD simulations may greatly reduce the force-field bias.


Assuntos
Entropia , Proteínas Intrinsicamente Desordenadas/química , Simulação de Dinâmica Molecular , Espalhamento a Baixo Ângulo , Difração de Raios X , Conformação Proteica
7.
J Chem Theory Comput ; 15(9): 5087-5102, 2019 Sep 10.
Artigo em Inglês | MEDLINE | ID: mdl-31402660

RESUMO

Circular dichroism (CD) spectroscopy is a highly sensitive but low-resolution technique to study the structure of proteins. Combined with molecular modeling or other complementary techniques, CD spectroscopy can provide essential information at higher resolution. To this end, we introduce a new computational method to calculate the electronic circular dichroism spectra of proteins from a structural model or ensemble using the average secondary structure composition and a precalculated set of basis spectra. The method is designed for model validation to estimate the error of a given protein structural model based on the measured CD spectrum. We compared the predictive power of our method to that of existing algorithms, namely, DichroCalc and PDB2CD, and found that it predicts CD spectra more accurately. Our results indicate that the derived basis sets are robust to both experimental errors in the reference spectra and the choice of the secondary structure classification algorithm. For over 80% of the globular reference proteins, our basis sets accurately predict the experimental spectrum solely from their secondary structure composition. For the remaining 20%, correcting for intensity normalization considerably improves the prediction power. Additionally, we show that the predictions for short peptides and an example complex of intrinsically disordered proteins strongly benefit from accounting for side-chain contributions and structural flexibility.


Assuntos
Dicroísmo Circular , Proteínas/química , Algoritmos , Modelos Moleculares , Conformação Proteica , Teoria Quântica
8.
J Chem Theory Comput ; 15(9): 5144-5153, 2019 Sep 10.
Artigo em Inglês | MEDLINE | ID: mdl-31411882

RESUMO

Nontargeted parallel cascade selection molecular dynamics (nt-PaCS-MD) is an enhanced conformational sampling method of proteins, which does not rely on knowledge of the target structure. It makes use of cyclic resampling from some relevant initial structures to expand the searched conformational subspace. The efficiency of nt-PaCS-MD depends on the selections of these initial structures. They are usually stochastically occurring perturbed structures at which larger conformation transitions are about to happen. Reliable identification of these is the key to using nt-PaCS-MD. Two new parameters, the moving root-mean-square deviation (mRMSD) and the inner products of the backbone dihedral angles Φ and Ψ, are introduced as indicators of conformational outliers in MD trajectories. Both are based on the analysis of a time-localized set of coordinates, overcoming the need for a target structure while still capturing the complexity of the conformational transition. The reference to which the mRMSD relates is the close surrounding of the i-th conformation, often the (i-1)st one. Hence the name "time-localized" analysis. In this work, we focus on its interplay with nt-PaCS-MD and show that it increases its effectiveness compared to older versions. The target system is the midsized protein T4 lysozyme (in explicit water) on which we demonstrate the open-closed transition without referring to any target configuration. Additionally, we show that the short MD trajectories can be used for the construction of a free energy landscape of the conformational transition based on the Markov state model.


Assuntos
Simulação de Dinâmica Molecular , Muramidase/química , Bacteriófago T4/enzimologia , Muramidase/metabolismo , Conformação Proteica , Fatores de Tempo
9.
J Chem Theory Comput ; 15(9): 5154-5160, 2019 Sep 10.
Artigo em Inglês | MEDLINE | ID: mdl-31412199

RESUMO

Side-chain modeling plays a critical role in protein structure prediction. However, in many current methods, balancing the speed and accuracy is still challenging. In this paper, on the basis of our previous work OPUS-Rota (Protein Sci. 2008, 17, 1576-1585), we introduce a new side-chain modeling method, OPUS-Rota2, which is tested on both a 65-protein test set (DB65) in the OPUS-Rota paper and a 379-protein test set (DB379) in the SCWRL4 paper. If the main chain is native, OPUS-Rota2 is more accurate than OPUS-Rota, SCWRL4, and OSCAR-star but slightly less accurate than OSCAR-o. Also, if the main chain is non-native, OPUS-Rota2 is more accurate than any other method. Moreover, OPUS-Rota2 is significantly faster than any other method, in particular, 2 orders of magnitude faster than OSCAR-o. Thus, the combination of higher accuracy and speed of OPUS-Rota2 in modeling side chains on both the native and non-native main chains makes OPUS-Rota2 a very useful tool in protein structure modeling.


Assuntos
Modelos Moleculares , Proteínas/química , Algoritmos , Interações Hidrofóbicas e Hidrofílicas , Conformação Proteica
10.
J Agric Food Chem ; 67(38): 10713-10725, 2019 Sep 25.
Artigo em Inglês | MEDLINE | ID: mdl-31453702

RESUMO

Converting peanut protein biomass waste into environmentally friendly meat substitutes by a high-moisture extrusion process can help solve both resource and waste problems and be "double green". A multiscale method combined with some emerging techniques such as atomic force microscopy-based infrared spectroscopy and X-ray microscopy was used to make the whole extrusion process visible to show the process of forming a meat-like fibrous structure using two-dimensional and three-dimensional perspectives. The results showed that the protein molecules underwent dramatic structural changes and unfolded in the extruder barrel, which created favorable conditions for molecular rearrangement in the subsequent zones. It was confirmed that the meat-like fibrous structure started to form at the junction of the die and the cooling zone and that this structure was caused by the phase separation and rearrangement of protein molecules in the cooling zone. Moreover, the interactions between hydrogen bonds and disulfide bonds formed in the cooling zone maintained the meat-like fibrous structure with an α-helix > ß-sheet > ß-turn > random coil. Of the two main peanut proteins, arachin played a greater role in forming the fibrous structure than conarachin, especially those subunits of arachin with a molecular weight of 42, 39, and 22 kDa.


Assuntos
Arachis/química , Aromatizantes/química , Extratos Vegetais/química , Proteínas de Plantas/química , Resíduos/análise , Aromatizantes/isolamento & purificação , Química Verde , Ligações de Hidrogênio , Raios Infravermelhos , Extratos Vegetais/isolamento & purificação , Conformação Proteica
11.
J Agric Food Chem ; 67(37): 10470-10480, 2019 Sep 18.
Artigo em Inglês | MEDLINE | ID: mdl-31469565

RESUMO

Foodborne nanoparticles (FNPs) produced by roasting have attracted the attention of people, owing to their safety risk to body health. Herein, we reported the formation, physicochemical properties, elemental composition, biodistribution, and binding with human serum albumin (HSA) of FNPs extracted from roast squid. The results showed that the FNP size gradually decreased from 4.1 to 2.3 nm as the roasting temperature changed from 190 to 250 °C. The main component elements of FNPs are carbon, oxygen, and nitrogen, and the carbon and nitrogen contents of FNPs increased with the roasting temperature rising. The surface of FNPs contained hydroxyl, amino, and carboxyl functional groups. The FNPs can emit fluorescence in ultraviolet light and show excitation-dependent emission behavior. Furthermore, it was found that the FNPs derived from roast squid could be accumulated in the stomach, intestine, and brain of BALB/c mice after oral feeding. Static fluorescence quenching of HSA was found by the Stern-Volmer equation and ultraviolet-visible spectrum analysis after interaction with the FNPs. After the addition of FNPs, the α-helix content of HSA decreased and the morphological height of HSA increased, which indicated that the FNPs could cause structural changes in HSA. The atomic force microscopy characterization showed the formation of nanocorona between FNPs and HSA.


Assuntos
Decapodiformes/química , Nanopartículas/química , Nanopartículas/metabolismo , Albumina Sérica Humana/química , Animais , Culinária , Decapodiformes/metabolismo , Fluorescência , Temperatura Alta , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Microscopia de Força Atômica , Tamanho da Partícula , Ligação Proteica , Conformação Proteica , Albumina Sérica Humana/metabolismo , Distribuição Tecidual
12.
Phys Chem Chem Phys ; 21(35): 18958-18969, 2019 Sep 21.
Artigo em Inglês | MEDLINE | ID: mdl-31453590

RESUMO

Enhanced sampling has been extensively used to capture the conformational transitions in protein folding, but it attracts much less attention in the studies of protein-protein recognition. In this study, we evaluated the impact of enhanced sampling methods and solute dielectric constants on the overall accuracy of the molecular mechanics/Poisson-Boltzmann surface area (MM/PBSA) and molecular mechanics/generalized Born surface area (MM/GBSA) approaches for the protein-protein binding free energy calculations. Here, two widely used enhanced sampling methods, including aMD and GaMD, and conventional molecular dynamics (cMD) simulations with two AMBER force fields (ff03 and ff14SB) were used to sample the conformations for 21 protein-protein complexes. The MM/PBSA and MM/GBSA calculation results illustrate that the standard MM/GBSA based on the cMD simulations yields the best Pearson correlation (rp = -0.523) between the predicted binding affinities and the experimental data, which is much higher than that given by MM/PBSA (rp = -0.212). Two enhanced sampling methods (aMD and GaMD) are indeed more efficient for conformational sampling, but they did not improve the binding affinity predictions for protein-protein systems, suggesting that the aMD or GaMD sampling (at least in short timescale simulations) may not be a good choice for the MM/PBSA and MM/GBSA predictions of protein-protein complexes. The solute dielectric constant of 1.0 is recommended to MM/GBSA, but a higher solute dielectric constant is recommended to MM/PBSA, especially for the systems with higher polarity on the protein-protein binding interfaces. Then, a preliminary assessment of the MM/GBSA calculations based on a variable dielectric generalized Born (VDGB) model was conducted. The results highlight the potential power of VDGB in the free energy predictions for protein-protein systems, but more thorough studies should be done in the future.


Assuntos
Técnicas de Química Analítica/métodos , Modelos Químicos , Proteínas/química , Técnicas de Química Analítica/normas , Simulação de Dinâmica Molecular , Ligação Proteica , Conformação Proteica , Reprodutibilidade dos Testes
13.
Phys Chem Chem Phys ; 21(33): 18149-18160, 2019 Aug 21.
Artigo em Inglês | MEDLINE | ID: mdl-31389436

RESUMO

Conformational entropies are of great interest when studying the binding of small ligands to proteins or the interaction of proteins. Unfortunately, there are no experimental methods available to measure conformational entropies of all groups in a protein. Instead, they are normally estimated from molecular dynamics (MD) simulations, although such methods show problems with convergence and correlation of motions, and depend on the accuracy of the underlying potential-energy function. Crystallographic atomic displacement parameters (also known as B-factors) are available in all crystal structures and contain information about the atomic fluctuations, which can be converted to entropies. We have studied whether B-factors can be employed to extract conformational entropies for proteins by comparing such entropies to those measured by NMR relaxation experiments or obtained from MD simulations in solution or in the crystal. Unfortunately, our results show that B-factor entropies are unreliable, because they include the movement and rotation of the entire protein, they exclude correlation of the movements and they include contributions other than the fluctuations, e.g. static disorder, as well as errors in the model and the scattering factors. We have tried to reduce the first problem by employing translation-libration-screw refinement, the second by employing a description of the correlated movement from MD simulations, and the third by studying only the change in entropy when a pair of ligands binds to the same protein, thoroughly re-refining the structures in exactly the same way and using the same set of alternative conformations. However, the experimental B-factors seem to be incompatible with fluctuations from MD simulations and the precision is too poor to give any reliable entropies.


Assuntos
Simulação de Dinâmica Molecular , Proteínas/química , Temperatura Ambiente , Cristalografia por Raios X , Entropia , Galectina 3/química , Ligantes , Muramidase/química , Conformação Proteica , Tripsina/química
14.
J Agric Food Chem ; 67(35): 9868-9876, 2019 Sep 04.
Artigo em Inglês | MEDLINE | ID: mdl-31389242

RESUMO

Amylosucrase (EC 2.4.1.4, ASase), a typical carbohydrate-active enzyme, can catalyze 5 types of reactions and recognize more than 50 types of glycosyl acceptors. However, most ASases are unstable even at 50 °C, which limits their practical industrial applications. In this study, an extremely thermostable ASase was discovered from Calidithermus timidus DSM 17022 (CT-ASase) with an optimal activity temperature of 55 °C, half-life of 1.09 h at 70 °C, and melting temperature of 74.47 °C. The recombinant CT-ASase was characterized as the first tetrameric ASase, and a structure-based truncation mutation was conducted to confirm the effect of tetrameric conformation on its thermostability. In addition, α-1,4-glucan was found to be the predominant product of CT-ASase at pH 6.0-8.0 and 30-60 °C.


Assuntos
Proteínas de Bactérias/química , Glucosiltransferases/química , Thermus/enzimologia , Sequência de Aminoácidos , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Estabilidade Enzimática , Glucosiltransferases/genética , Glucosiltransferases/metabolismo , Temperatura Alta , Concentração de Íons de Hidrogênio , Cinética , Conformação Proteica , Alinhamento de Sequência , Thermus/química , Thermus/genética
15.
BMC Bioinformatics ; 20(1): 419, 2019 Aug 13.
Artigo em Inglês | MEDLINE | ID: mdl-31409275

RESUMO

BACKGROUND: Alignment of sequence families described by profiles provides a sensitive means for establishing homology between proteins and is important in protein evolutionary, structural, and functional studies. In the context of a steadily growing amount of sequence data, estimating the statistical significance of alignments, including profile-profile alignments, plays a key role in alignment-based homology search algorithms. Still, it is an open question as to what and whether one type of distribution governs profile-profile alignment score, especially when profile-profile substitution scores involve such terms as secondary structure predictions. RESULTS: This study presents a methodology for estimating the statistical significance of this type of alignments. The methodology rests on a new algorithm developed for generating random profiles such that their alignment scores are distributed similarly to those obtained for real unrelated profiles. We show that improvements in statistical accuracy and sensitivity and high-quality alignment rate result from statistically characterizing alignments by establishing the dependence of statistical parameters on various measures associated with both individual and pairwise profile characteristics. Implemented in the COMER software, the proposed methodology yielded an increase of up to 34.2% in the number of true positives and up to 61.8% in the number of high-quality alignments with respect to the previous version of the COMER method. CONCLUSIONS: The more accurate estimation of statistical significance is implemented in the COMER method, which is now more sensitive and provides an increased rate of high-quality profile-profile alignments. The results of the present study also suggest directions for future research.


Assuntos
Modelos Teóricos , Proteínas/química , Algoritmos , Sequência de Aminoácidos , Conformação Proteica , Alinhamento de Sequência
16.
Phys Rev Lett ; 122(23): 238102, 2019 Jun 14.
Artigo em Inglês | MEDLINE | ID: mdl-31298900

RESUMO

The enormous catalytic power of natural enzymes relies on the ability to overcome the bottleneck event in the enzymatic cycle, yet the underlying physical mechanisms are not fully understood. Here, by performing molecular simulations of the whole enzymatic cycle for a model multisubstrate enzyme with a dynamic energy landscape model, we show that multisubstrate enzymes can utilize steric frustration to facilitate the rate-limiting product-release step. During the enzymatic cycles, the bottleneck product is actively squeezed out by the binding of a new substrate at the neighboring site through the population of a substrate-product cobound complex, in which the binding pockets are frustrated due to steric incompatibility. Such steric frustration thereby enables an active mechanism of product release driven by substrate-binding energy, facilitating the enzymatic cycle.


Assuntos
Enzimas/química , Enzimas/metabolismo , Modelos Químicos , Adenilato Quinase/química , Adenilato Quinase/metabolismo , Modelos Moleculares , Conformação Proteica , Relação Estrutura-Atividade , Termodinâmica
17.
Inorg Chem ; 58(15): 9672-9690, 2019 Aug 05.
Artigo em Inglês | MEDLINE | ID: mdl-31282663

RESUMO

We have studied the geometry and electronic structure of the P-cluster in nitrogenase in four oxidation states: PN, P1+, P2+, and P3+. We have employed combined quantum mechanical and molecular mechanical (QM/MM) calculations, using two different density-functional theory methods, TPSS and B3LYP. The calculations confirm that the side chain of Ser-188 is most likely deprotonated in the partly oxidized P1+ state, thereby forming a bond to Fe6. Likewise, the backbone amide group of Cys-88 is deprotonated in the doubly oxidized P2+ state, forming a bond to Fe5. The calculations also confirm the two conformations of the P-cluster in the atomic-resolution crystal structure of the enzyme, representing the PN and P2+ states, but show that the finer differences between the two structures are not fully reflected in the crystal structure, because the coordinates of only two atoms differ between the two conformations. However, the recent crystal structure of the P1+ state seems to be of lower quality with many dubious Fe-Fe and Fe-S distances. Quantum refinement of this structure indicates that it is a mixture of the P1+ and P2+ states but confirms that the side chain of Ser-188 is most likely deprotonated in both states. TPSS gives structures that are appreciably closer to the crystal structures than does B3LYP. In addition, we have studied all 16-48 possible broken-symmetry states of the four oxidation states of the P-cluster with DFT in the one or two observed spin states. For the reduced PN state, we can settle the most likely state from the calculated energies and geometries. However, for the more oxidized states there are large differences in the predictions obtained with the two DFT methods.


Assuntos
Nitrogenase/química , Teoria Quântica , Elétrons , Modelos Moleculares , Nitrogenase/metabolismo , Oxirredução , Conformação Proteica
18.
J Chem Theory Comput ; 15(8): 4699-4707, 2019 Aug 13.
Artigo em Inglês | MEDLINE | ID: mdl-31314523

RESUMO

We present a fast implementation of the nudged elastic band (NEB) method into the particle mesh Ewald molecular dynamics module of the Amber software package for both central processing units (CPU) and graphics processing units (GPU). The accuracy of the new implementation has been validated for three cases: a conformational change of alanine dipeptide, the α-helix to ß-sheet transition in polyalanine, and a large conformational transition in the human 8-oxoguanine-DNA glycosylase with DNA complex (OGG1-DNA). Timing benchmark tests were performed on the explicitly solvated OGG1-DNA system containing ∼50 000 atoms. The GPU-optimized implementation of NEB achieves a more than two orders of magnitude speedup compared with the previous CPU implementation performed with a two-core CPU processor. The speed and scalable features of this implementation will enable NEB applications on larger and more complex systems.


Assuntos
DNA Glicosilases/química , Dipeptídeos/química , Simulação de Dinâmica Molecular , Peptídeos/química , Gráficos por Computador , DNA/química , Humanos , Conformação Proteica , Conformação Proteica em alfa-Hélice , Conformação Proteica em Folha beta , Software , Fatores de Tempo
19.
Phys Chem Chem Phys ; 21(28): 15686-15694, 2019 Jul 17.
Artigo em Inglês | MEDLINE | ID: mdl-31271401

RESUMO

Alzheimer's disease is associated with the abnormal self-assembly of amyloid-ß (Aß) peptide into toxic oligomers and fibrils. Recent experiments reported that Aß16-22, containing the central hydrophobic core (CHC) of Aß, formed antiparallel ß-sheet fibrils, while its E22Q mutant self-assembled into parallel ß-sheet fibrils. However, the molecular mechanisms underlying E22Q-mutation-induced parallel ß-sheet fibril formation are not well understood. Herein, we performed molecular dynamics (MD) simulations to study the dimerization processes of Aß16-22 and Aß16-22E22Q peptides. ß-Sheet dimers with diverse hydrogen bond arrangements were observed and they exhibited highly dynamic and interconverting properties. An antiparallel-to-parallel ß-sheet transition occurred in the assembly process of the E22Q mutant, but not in that of Aß16-22. During this conformational transformation process, the inter-molecular Q22-Q22 hydrogen bonds were first formed and acted as a binder to facilitate the two chains forming a parallel orientation, then the hydrophobic interactions between residues in the CHC region consolidated this arrangement and drove the main-chain H-bond formation, hence resulting in parallel ß-sheet formation. However, parallel ß-sheets were less populated than antiparallel ß-sheets of Aß16-22E22Q dimers. In order to explore whether parallel ß-sheets became dominant in larger size oligomers, we investigated the conformational ensembles of Aß16-22 and Aß16-22E22Q octamers by conducting replica exchange molecular dynamics (REMD) simulations. The REMD simulations revealed that the population of parallel ß-strand alignment increased with an increase of the size of ordered Aß16-22E22Q ß-sheet oligomers, implying that the formation of full parallel ß-sheets requires larger sized oligomers. Our findings provide a mechanistic explanation for the E22Q-mutation-induced formation of parallel ß-sheet fibrils observed experimentally.


Assuntos
Simulação de Dinâmica Molecular , Mutação , Conformação Proteica em Folha beta/genética , Amiloide/química , Peptídeos beta-Amiloides/química , Peptídeos beta-Amiloides/genética , Humanos , Conformação Proteica
20.
Arch Virol ; 164(9): 2297-2307, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31267215

RESUMO

The influence of the high genetic variability of hepatitis B virus (HBV) on the sensitivity of serological assays has received little attention so far. A major source of variability is related to viral genotypes and subgenotypes. Their possible influence on diagnosis and prophylaxis is poorly known and has mostly been evaluated for genotypes A, B, C and D. Robust data showing the detection efficiency of HBsAg from genotype F is lacking. This study examined the effect of virus-like particles containing HBsAg from genotypes A and F (particularly, F1b and F4) produced in Pichia pastoris in relation to the anti-HBs antibodies used in the immunoassays for in vitro diagnosis and compared it with that exerted by the G145R S-escape mutant. The results showed that HBsAg detection rates for subgenotypes F1b and F4 differed significantly from those obtained for genotype A and that subgenotype F1b had a major impact on the sensitivity of the immunoassays tested. Prediction of the tertiary structure of subgenotypes F1b and F4 revealed changes inside and outside the major hydrophilic region (aa 101-160) of the HBsAg compared to genotype A and the G145R variant. A phosphorylation site (target for protein kinase C) produced by the G145R substitution might prevent recognition by anti-HBs antibodies. In conclusion, the use of different genotypes or variants for diagnosis could improve the rate of detection of HBV infection. The incorporation of a genotype-F-derived HBsAg vaccine in areas where this genotype is endemic should be evaluated, since this might also affect vaccination efficacy.


Assuntos
Antígenos de Superfície da Hepatite B/genética , Vírus da Hepatite B/isolamento & purificação , Hepatite B/virologia , Sequência de Aminoácidos , Hepatite B/diagnóstico , Antígenos de Superfície da Hepatite B/química , Antígenos de Superfície da Hepatite B/metabolismo , Vírus da Hepatite B/química , Vírus da Hepatite B/classificação , Vírus da Hepatite B/genética , Humanos , Dados de Sequência Molecular , Mutação , Filogenia , Conformação Proteica , Alinhamento de Sequência
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