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1.
Phys Chem Chem Phys ; 21(44): 24601-24619, 2019 Nov 13.
Artigo em Inglês | MEDLINE | ID: mdl-31670335

RESUMO

Natural bond orbital (NBO) analysis of electron delocalization in a series of capped isolated peptides is used to diagnose amide-amide H-bonding and backbone-induced hyperconjugative interactions, and to rationalize their spectral effects. The sum of the stabilization energies corresponding to the interactions between NBOs that are involved in the H-bonding is demonstrated as an insightful indicator for the H-bond strength. It is then used to decouple the effect of the H-bond distance from that, intrinsic, of the donor/acceptor relative orientation, i.e., the geometrical approach. The diversity of the approaches given by the series of peptides studied enables us to illustrate the crucial importance of the approach when the acceptor is a carbonyl group, and emphasizes that efficient approaches can be achieved despite not matching the usual picture of a proton donor directly facing a lone pair of the proton acceptor, i.e., that encountered in intermolecular H-bonds. The study also illustrates the role of backbone flexibility, partly controlled by backbone-amide hyperconjugative interactions, in influencing the equilibrium structures, in particular by frustrating or enhancing the HB for a given geometrical approach. Finally, the presently used NBO-based HB strength indicator enables a fair prediction of the frequency of the proton donor amide NH stretching mode, but this simple picture is blurred by ubiquitous hyperconjugative effects between the backbone and amide groups, whose magnitude can be comparable to that of the weakest H-bonds.


Assuntos
Amidas/química , Peptídeos/química , Dimerização , Ligações de Hidrogênio , Modelos Moleculares , Conformação Proteica em alfa-Hélice , Conformação Proteica em Folha beta , Teoria Quântica
2.
Chem Commun (Camb) ; 55(83): 12463-12466, 2019 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-31576854

RESUMO

Herein, an ultrasensitive electrochemical biosensor is proposed for the quantification of the Flu A virus biomarker DNA (fDNA), and is based on loop-mediated isothermal amplification-generated hydrogen ions (LAMP-H+) which induce the formation of the dimer i-motif structure (DiMS) for signal transduction, coupled with exonuclease III (ExoIII)-assisted DNA walking for signal dual-amplification.


Assuntos
Técnicas Biossensoriais , DNA Viral/análise , Técnicas Eletroquímicas , Vírus da Influenza A/química , Técnicas de Amplificação de Ácido Nucleico , Prótons , Biomarcadores/análise , Dimerização , Íons/química
3.
Phys Chem Chem Phys ; 21(41): 22763-22773, 2019 Oct 24.
Artigo em Inglês | MEDLINE | ID: mdl-31595279

RESUMO

The adenosine A1 receptor (A1R) is one of four adenosine receptors in humans, which are involved in the function of the cardiovascular, respiratory and central nervous systems. Experimental results indicate that A1R can form a homodimer and that the protomer-protomer interaction in the A1R dimer is related to certain pharmacological characteristics of A1R activation. In this work, we performed docking, metadynamics simulation, conventional molecular dynamics simulations, Gaussian-accelerated molecular dynamics simulations, potential of mean force calculations, dynamic cross-correlation motions analysis and community network analysis to study the binding mode of 5'-N-ethylcarboxamidoadenosine (NECA) to A1R and the effect of dimerization on the activation of A1R. Our results show that NECA binds to A1R in a similar mode to adenosine in the A1R crystal structure and NECA in the A2AR crystal structure. The A1R homodimer can be activated by one or two agonists with NECA occupying its orthosteric pockets in one (which we call the NECA-A1R system) or both protomers (which we call the dNECA-A1R system). In the NECA-A1R system, activation is predicated in the protomer without NECA bound. In the dNECA-A1R system, only one protomer achieves the active state. These findings suggest an asymmetrical activation mechanism of the homodimer and a negative cooperativity between the two protomers. We envision that our results may further facilitate the drug development of A1R.


Assuntos
Dimerização , Modelos Moleculares , Receptor A1 de Adenosina/química , Receptor A1 de Adenosina/metabolismo , Cristalização , Humanos , Simulação de Dinâmica Molecular , Ligação Proteica , Estrutura Terciária de Proteína
4.
J Chem Theory Comput ; 15(11): 6444-6455, 2019 Nov 12.
Artigo em Inglês | MEDLINE | ID: mdl-31593632

RESUMO

Integral membrane proteins are ubiquitous in biological cellular and subcellular membranes. Despite their significance to cell function, isolation of membrane proteins from their hydrophobic lipid environment and further characterization remains a challenge. To obtain insights into membrane proteins, computational approaches such as docking or self-assembly simulations have been used; however, the promise of these approaches has been limited due to the computational cost. Here we present a new approach called Protein AssociatioN Energy Landscape (PANEL) that provides an extensive and converged data set for all possible conformations of membrane protein associations using a combination of stochastic sampling and equilibration simulations. The PANEL method samples the rotational space around both interacting proteins to obtain the comprehensive interaction energy landscape. We demonstrate the versatility of the PANEL method using two distinct applications: (a) dimerization of claudin-5 tight junction proteins in phospholipid bilayer membrane and (b) dimer and trimer formation of the Outer membrane protein F (OmpF) in the lipopolysaccharide-rich bacterial outer membrane. Both applications required only a fraction of simulation cost compared to self-assembly simulations. The method is robust as it can capture changes in protein-protein conformations caused by point mutations. Moreover, the method is versatile and independent of the molecular resolution (atomistic or coarse grain) or the choice of force field employed to compute the pair-interaction energies. The PANEL method is implemented in easy-to-use scripts that are available for download for general use by the scientific community to characterize any pair of interacting integral membrane proteins.


Assuntos
Proteínas de Membrana/química , Modelos Moleculares , Bactérias/metabolismo , Membrana Celular/química , Membrana Celular/metabolismo , Dimerização , Proteínas de Membrana/metabolismo , Mutação Puntual , Porinas/química , Porinas/genética , Porinas/metabolismo , Conformação Proteica , Termodinâmica
5.
J Agric Food Chem ; 67(41): 11498-11507, 2019 Oct 16.
Artigo em Inglês | MEDLINE | ID: mdl-31544455

RESUMO

Proanthocyanidins (PAs) possess superior antioxidant properties and nutritious value, however, low bioavailability and stability limit their applications. Here, we developed a novel method to encapsulate PA dimers successfully into horse spleen apoferritin (apoHSF) using a disassembly/reassembly method based on pH change. The PA-HSF nanoparticles were characterized using fluorescence spectroscopy, transmission electron microscopy, circular dichroism, and high-performance liquid chromatography. One apoferritin cage could approximately encapsulate 25.6 molecules of the PA dimer. The results showed that the encapsulation of the PA dimers protected it from the damage of oxidants and temperature below room temperature would be an appropriate condition for HSF-578 solution storage. Moreover, HepG2 cell monolayer absorption and adhesion analyses indicated that the PA dimers encapsulated within apoHSF cages were more efficient in transport. In addition, it was indicated that the PA-HSF nanoparticles had higher cellular antioxidant activity. The novel strategy provided in this study indicates that the protein cage structures like ferritin have potential to be applied in the field of food nutrition.


Assuntos
Antioxidantes/química , Antioxidantes/metabolismo , Apoferritinas/química , Composição de Medicamentos/métodos , Proantocianidinas/química , Proantocianidinas/metabolismo , Animais , Disponibilidade Biológica , Transporte Biológico , Cromatografia Líquida de Alta Pressão , Dimerização , Células Hep G2 , Cavalos , Humanos
6.
Chem Commun (Camb) ; 55(78): 11671-11674, 2019 Sep 26.
Artigo em Inglês | MEDLINE | ID: mdl-31497827

RESUMO

We report the design and optimisation of novel oligonucleotide substrates for a sensitive fluorescence assay for high-throughput screening and functional studies of the DNA repair enzyme, XPF-ERCC1, with a view to accelerating inhibitor and drug discovery.


Assuntos
Reparo do DNA , Proteínas de Ligação a DNA/metabolismo , Endonucleases/metabolismo , Proteínas de Ligação a DNA/química , Proteínas de Ligação a DNA/genética , Dimerização , Endonucleases/química , Endonucleases/genética , Humanos , Oligonucleotídeos/química , Oligonucleotídeos/metabolismo , Especificidade por Substrato , Temperatura Ambiente
7.
J Chem Theory Comput ; 15(11): 6403-6410, 2019 Nov 12.
Artigo em Inglês | MEDLINE | ID: mdl-31525924

RESUMO

Interaction of the small ubiquitin-related modifier (SUMO) and peptides containing a SUMO-interacting motif (SIM) has attracted a lot of interest in recent years, yet their structural properties and relationships between the composition of the peptide and binding free energy are not completely understood. We perform molecular dynamics simulations of the complex formed by SUMO and a peptide containing the tight-binding SIM of the protein inhibitor of activated STAT. The calculated standard binding free energy of -5.06 kcal/mol is in reasonable agreement with the experimental value of -6.54 kcal/mol. Experimental results for complexes formed by SUMO and SIM dimers indicate the existence of a parallel and an antiparallel binding mode for similar SIM peptides. We find that the parallel binding mode is highly favored in the present case. Furthermore, the simulations show that residues neighboring the SIM core motif contribute strongly to the binding energy. Structurally, the complex shows differences from the picture in which the SIM core motif lies deep in the SUMO binding groove. This also supports the idea that neighboring residues play an important role in binding.


Assuntos
Peptídeos/química , Proteínas Modificadoras Pequenas Relacionadas à Ubiquitina/química , Motivos de Aminoácidos , Dimerização , Simulação de Dinâmica Molecular , Peptídeos/metabolismo , Ligação Proteica , Proteínas Modificadoras Pequenas Relacionadas à Ubiquitina/metabolismo , Termodinâmica
8.
J Chem Theory Comput ; 15(11): 5894-5907, 2019 Nov 12.
Artigo em Inglês | MEDLINE | ID: mdl-31538779

RESUMO

A nonempirical quantum mechanical method for the efficient and accurate quantification and analysis of intermolecular interactions is presented and tested on existing benchmark sets. The leading idea here is to focus on the intermolecular part of the correlation energy that contains the all-important London dispersion (LD) interaction. To keep the cost of the method low, essentially at the level of a Hartree-Fock (HF) calculation, the intramolecular part of the correlation energy is neglected. We also neglect the nondispersive parts of the intermolecular correlation energy. This scheme that we denote as Hartree-Fock plus London dispersion (HFLD) can be readily realized on the basis of the recently reported multilevel implementation of the domain-based local pair natural orbital coupled-cluster (DLPNO-CC) theory in conjunction with the well-established local energy decomposition (LED) analysis. The accuracy and efficiency of the HFLD method are evaluated on rare gas dimers, on the S66 and L7 benchmark sets of noncovalent interactions, and on an additional set (LP14) consisting of bulky Lewis pairs held together by intermolecular interactions of various strengths, with interaction energies ranging from -8 to -107 kcal/mol. It is first shown that the LD energy calculated with this approach is essentially identical to that obtained from the full DLPNO-CCSD(T)/LED calculation, with a mean absolute error of 0.2 kcal/mol on the S66 benchmark set. Moreover, in terms of the overall interaction energies, the HFLD method shows an efficiency that is comparable to that of the HF method, while retaining an accuracy between that of the DLPNO-CCSD and DLPNO-CCSD(T) schemes. Since the underlying DLPNO-CCSD method is linear scaling with respect to the system size, the HFLD approach also does not lead to new bottlenecks for large systems. As an illustrative example of its efficiency, the HFLD scheme was applied to the interaction between the substrate and the residues in the active site of the cyclohexanone monooxygenase enzyme. The excellent cost/performance ratio indicates that the HFLD method opens new avenues for the accurate calculation and analysis of noncovalent interaction energies in large molecular systems.


Assuntos
Modelos Moleculares , Teoria Quântica , Domínio Catalítico , Dimerização , Gases/química , Oxigenases/química , Oxigenases/metabolismo , Especificidade por Substrato , Termodinâmica
9.
J Chem Theory Comput ; 15(10): 5659-5673, 2019 Oct 08.
Artigo em Inglês | MEDLINE | ID: mdl-31476125

RESUMO

Human heterochromatin protein 1 (HP1) is a key factor in heterochromatin formation and maintenance. Its chromo-shadow domain (CSD) homodimerizes, and the HP1 dimer acts as a hub, transiently interacting with diverse binding partner (BP) proteins. We analyze atomistic details of interactions of the HP1γ(CSD) dimer with one of its targets, the histone H3 N-terminal tail, using molecular dynamics (MD) simulations. The goal is to complement the available X-ray crystallography data and unravel potential dynamic effects in the molecular recognition. Our results suggest that HP1(CSD)-BP recognition involves structural dynamics of both partners, including structural communication between adjacent binding pockets that may fine-tune the sequence recognition. For example, HP1 Trp174 sidechain substates may help in distinguishing residues bound in the conserved HP1(CSD) ±2 hydrophobic pockets. Further, there is intricate competition between the binding of negatively charged HP1 C-terminal extension and solvent anions near the ±2 hydrophobic pockets, which is also influenced by the BP sequence. Phosphorylated H3 Y41 can interact with the same site. We also analyze the ability of several pair-additive force fields to describe the protein-protein interface. ff14SB and ff99SB-ILDN* provide the closest correspondence with the crystallographic model. The ff15ipq local dynamics are somewhat less consistent with details of the experimental structure, while larger perturbations of the interface commonly occur in CHARMM36m simulations. The balance of some interactions, mainly around the anion binding site, also depends on the ion parameters. Some differences between the simulated and experimental structures are attributable to crystal packing.


Assuntos
Proteínas Cromossômicas não Histona/química , Histonas/química , Cristalografia por Raios X , Dimerização , Histonas/metabolismo , Humanos , Simulação de Dinâmica Molecular , Conformação Proteica
10.
J Chem Theory Comput ; 15(10): 5448-5460, 2019 Oct 08.
Artigo em Inglês | MEDLINE | ID: mdl-31498621

RESUMO

The computational and conceptual simplifications realized by coarse-grain (CG) models make them a ubiquitous tool in the current computational modeling landscape. Building block based CG models, such as the Martini model, possess the key advantage of allowing for a broad range of applications without the need to reparametrize the force field each time. However, there are certain inherent limitations to this approach, which we investigate in detail in this work. We first study the consequences of the absence of specific cross Lennard-Jones parameters between different particle sizes. We show that this lack may lead to artificially high free energy barriers in dimerization profiles. We then look at the effect of deviating too far from the standard bonded parameters, both in terms of solute partitioning behavior and solvent properties. Moreover, we show that too weak bonded force constants entail the risk of artificially inducing clustering, which has to be taken into account when designing elastic network models for proteins. These results have implications for the current use of the Martini CG model and provide clear directions for the reparametrization of the Martini model. Moreover, our findings are generally relevant for the parametrization of any other building block based force field.


Assuntos
Simulação de Dinâmica Molecular , Dimerização , Tamanho da Partícula , Termodinâmica
11.
J Phys Chem Lett ; 10(19): 5861-5867, 2019 Oct 03.
Artigo em Inglês | MEDLINE | ID: mdl-31464127

RESUMO

Circularly polarized luminescence (CPL) reflects the excited-state properties of the chiral system. However, compared to the singlet and triplet excited states, there are still many unknowns about CPL from the double excited state. Here, using the self-assembly strategy of a dipeptide substituted naphthalenediimide (NDI-GE) and the photogenerated radical anions, we have explored the ground-state (CD) and excited-state (CPL) chiral characteristics of neutral NDI and NDI•- radical anion assemblies. The neutral gelator assemblies showed CPL with the dissymmetry factor glum on the order of 10-3; the radical anion exhibited an inversed CPL signal with a significantly enhanced glum of 10-1. Time-dependent density functional theory calculation revealed that upon formation of the radical anions, the direction of the dipole moment changed, thus leading to the inversion of CD and CPL. The present work reveals a new platform for developing CPL materials based on the doublet excited state.


Assuntos
Dipeptídeos/química , Imidas/química , Naftalenos/química , Ânions/química , Teoria da Densidade Funcional , Dimerização , Cinética , Luminescência , Modelos Químicos , Modelos Moleculares , Processos Fotoquímicos , Solventes/química
12.
Pharm Res ; 36(10): 150, 2019 Aug 19.
Artigo em Inglês | MEDLINE | ID: mdl-31428879

RESUMO

PURPOSE: The unconventional tabletability of the indomethacin polymorphs - α and γ - are investigated from a topological and mechanical perspective using powder Brillouin light scattering (p-BLS) to identify the specific structure-performance relationship in these materials. METHOD: Indomethacin (γ-form) was purchased and used to prepare the α polymorph. Powder X-ray diffraction was used to confirm phase identity, while p-BLS was used to obtain the mechanical properties. Energy frameworks were determined with Crystal Explorer to visualize the interaction topologies. Using a Carver press and a stress-strain analyzer, the tableting performance of each polymorph was determined. RESULTS: Polymorph-specific acoustic frequency distributions were observed with distinct, zero-porosity, aggregate elastic moduli determined. The p-BLS spectra for α-indomethacin display a population of low-velocity shear modes, indicating a direction of facilitated shear. This improves slip-mediated plasticity and tabletability. Our p-BLS spectra experimentally indicates that a low-energy slip system is available to α-indomethacin which supports ours and previous energy framework calculations. Despite a 2d-layered crystal motif favorable for shear deformation, the γ-form displays a higher shear modulus that is supported by our hydrogen-bonding analysis of γ-indomethacin. CONCLUSION: Our experimental, mechanical data is consistent with the predicted interaction topologies and these two inputs combined permit a comprehensive, molecular understanding of polymorph-specific tabletability.


Assuntos
Indometacina/química , Cristalização , Dimerização , Composição de Medicamentos , Ligações de Hidrogênio , Luz , Fenômenos Mecânicos , Porosidade , Pós , Espalhamento de Radiação , Comprimidos , Termodinâmica
13.
Chemistry ; 25(52): 12137-12144, 2019 Sep 18.
Artigo em Inglês | MEDLINE | ID: mdl-31276238

RESUMO

Isomerism heavily influences the optoelectronic properties and self-assembly behavior of compounds and subsequently affects their device performance. Herein, two pairs of isomeric perylene diimide (PDI) dimers, PDI and PDI2, were designed and synthesized. The electron-deficient 9,10-anthraquinone group was employed as the bridge, and thus, the resultant dimers exhibited an acceptor-acceptor-acceptor (A-A-A) structure. To determine the isomeric effects on the optoelectronic properties and photovoltaic performance of these dimers, their absorptivity, luminescence, and redox behavior were studied. Bulk heterojunction organic solar cells based on these four dimers were fabricated and measured. The two PDI dimers exhibited clear differences in photovoltaic performance, whereas the two PDI2 analogues showed similar power conversion efficiencies (PCEs). The PCEs of the two PDI2 dimers are much higher than those of the PDI dimers. These results illustrate that the isomeric effect of PDI dimers is much larger than that of PDI2 dimers on the device performance, and proper expansion of conjugation could improve the device performance.


Assuntos
Antraquinonas/química , Imidas/química , Perileno/análogos & derivados , Dimerização , Eletrônica , Isomerismo , Oxirredução , Perileno/química , Energia Solar
14.
J Photochem Photobiol B ; 198: 111546, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31351309

RESUMO

Fluorescence dynamics of human d-amino acid oxidase (hDAAO) and its five inhibitors have been studied in the picoseconds time domain, and compared with one in d-amino acid oxidase from porcine kidney (pkDAAO) reported. The fluorescence lifetimes were identified as 47 ps in the dimer, 235 ps in the monomer, which are compared with those of pkDAAO (45 ps-185 ps). The fluorescence lifetimes of the hDAAO did not change upon the inhibitor bindings despite of modifications in the absorption spectra. This indicates that the lifetimes of the complexes are too short to detect with the picosecond lifetime instrument. Numbers of the aromatic amino acids are similar between the both DAAOs. The fluorescence lifetimes of hDAAO were analysed with an ET theory using the crystal structure. The difference in the lifetimes of the dimer and monomer was well described in terms of difference in the electron affinity of the excited isoalloxazine (Iso*) between the two forms of the protein, though it is not known whether the structure of the monomer is different from the dimer. Three fastest ET donors were Tyr314, Trp52 and Tyr224 in the dimer, while Tyr314, Tyr224 and Tyr55 in the monomer, which are compared to those in pkDAAO, Tyr314, Tyr224 and Tyr228 in the dimer, and Tyr224, Tyr314 and Tyr228 in the monomer. The ET rate from Trp55 in hDAAO dimer was much faster compared to the rate in pkDAAO dimer. A rise component with negative pre-exponential factor was not observed in hDAAO, which are found in pkDAAO.


Assuntos
Aminoácidos Aromáticos/química , D-Aminoácido Oxidase/química , Flavinas/química , Rim/enzimologia , Animais , D-Aminoácido Oxidase/metabolismo , Dimerização , Transporte de Elétrons , Humanos , Ligações de Hidrogênio , Espectrometria de Fluorescência , Eletricidade Estática , Suínos
15.
Chem Biol Interact ; 310: 108756, 2019 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-31325422

RESUMO

Human butyrylcholinesterase (BChE) is a widely distributed plasma enzyme. For decades, numerous research efforts have been directed at engineering BChE as a bioscavenger of organophosphorus insecticides and chemical warfare nerve agents. However, it has been a grand challenge to cost-efficiently produce BChE in large-scale. Recently reported studies have successfully designed a truncated BChE mutant (with amino-acid substitutions on 47 residues that are far away from the catalytic site), denoted as BChE-M47 for convenience, which can be expressed in E. coli without loss of its catalytic activity. In this study, we aimed to dimerize the truncated BChE mutant protein expressed in a prokaryotic system (E. coli) in order to further improve its thermal stability by introducing a pair of cross-subunit disulfide bonds to the BChE-M47 structure. Specifically, the E377C/A516C mutations were designed and introduced to BChE-M47, and the obtained new protein entity, denoted as BChE-M48, with a pair of cross-subunit disulfide bonds indeed exists as a dimer with significantly improved thermostability and unaltered catalytic activity and reactivity compared to BChE-M47. These results provide a new strategy for optimizing protein stability for production in a cost-efficient prokaryotic system. Our enzyme, BChE-M48, has a half-life of almost one week at a 37°C, suggesting that it could be utilized as a highly stable bioscavenger of OP insecticides and chemical warfare nerve agents.


Assuntos
Butirilcolinesterase/metabolismo , Engenharia de Proteínas/métodos , Butirilcolinesterase/genética , Substâncias para a Guerra Química/metabolismo , Dimerização , Estabilidade Enzimática , Escherichia coli/genética , Humanos , Inseticidas/metabolismo , Proteínas Mutantes/genética , Proteínas Mutantes/metabolismo , Compostos Organofosforados/metabolismo
16.
Chem Commun (Camb) ; 55(66): 9769-9772, 2019 Aug 13.
Artigo em Inglês | MEDLINE | ID: mdl-31329196

RESUMO

We report the accelerated photoisomerization of amphiphilic lophine dimers based on the inner environments of molecular assemblies and rapid control of the interfacial properties of aqueous solution with photoirradiation. This novel photoisomerization system enables on-demand controlled release of drugs, perfumes, and other active compounds.


Assuntos
Imidazóis/química , Dimerização , Sistemas de Liberação de Medicamentos , Isomerismo , Processos Fotoquímicos , Espectrofotometria Ultravioleta , Tensão Superficial
17.
Biophys Chem ; 253: 106221, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31306918

RESUMO

The Ca2+ induced Spontaneous Oscillatory Contraction (Ca-SPOC) of cardiac myofibrils oscillate with a period similar to resting heartbeat of several animal species, and its auto-oscillatory properties set the basic rhythm of cardiac contraction. To explain the dynamics of Ca-SPOC, the present paper constructs a novel chemical kinetical model based upon the cooperative behavior between the two heads of myosin II dimer, also considering the reaction-diffusion effect of ATP inside myocardial fibers. The simulation results show that the concentration of ATP inside myocardial fibers oscillates over time under some special conditions, together with the proportions of myosin II dimers in different states periodically changing with time, which contributes to produce the sustained oscillations of contractive tension. These results indicate that the SPOC of muscles may be partly due to chemical oscillation involved in the actomyosin ATPase cycle, which has been ignored by the previous theoretical studies.


Assuntos
Cálcio/farmacologia , Modelos Biológicos , Contração Miocárdica/efeitos dos fármacos , Miocárdio/metabolismo , Trifosfato de Adenosina/metabolismo , Sinalização do Cálcio/efeitos dos fármacos , Dimerização , Cinética , Miosina Tipo II/metabolismo
18.
Nat Cell Biol ; 21(7): 867-878, 2019 07.
Artigo em Inglês | MEDLINE | ID: mdl-31263268

RESUMO

mTORC2 plays critical roles in metabolism, cell survival and actin cytoskeletal dynamics through the phosphorylation of AKT. Despite its importance to biology and medicine, it is unclear how mTORC2-mediated AKT phosphorylation is controlled. Here, we identify an unforeseen principle by which a GDP-bound form of the conserved small G protein Rho GTPase directly activates mTORC2 in AKT phosphorylation in social amoebae (Dictyostelium discoideum) cells. Using biochemical reconstitution with purified proteins, we demonstrate that Rho-GDP promotes AKT phosphorylation by assembling a supercomplex with Ras-GTP and mTORC2. This supercomplex formation is controlled by the chemoattractant-induced phosphorylation of Rho-GDP at S192 by GSK-3. Furthermore, Rho-GDP rescues defects in both mTORC2-mediated AKT phosphorylation and directed cell migration in Rho-null cells in a manner dependent on phosphorylation of S192. Thus, in contrast to the prevailing view that the GDP-bound forms of G proteins are inactive, our study reveals that mTORC2-AKT signalling is activated by Rho-GDP.


Assuntos
Movimento Celular/fisiologia , Dimerização , Alvo Mecanístico do Complexo 2 de Rapamicina/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Animais , Citoesqueleto/metabolismo , Proteínas de Ligação ao GTP/metabolismo , Quinase 3 da Glicogênio Sintase/metabolismo , Guanosina Difosfato/metabolismo , Humanos , Fosforilação/fisiologia
19.
J Chem Ecol ; 45(5-6): 490-501, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31175497

RESUMO

The development of pesticide resistance in insects and recent bans on pesticides call for the identification of natural sources of resistance in crops. Here, we used natural variation in pepper (Capsicum spp.) resistance combined with an untargeted metabolomics approach to detect secondary metabolites related to thrips (Frankliniella occidentalis) resistance. Using leaf disc choice assays, we tested 11 Capsicum accessions of C. annuum and C. chinense in both vegetative and flowering stages for thrips resistance. Metabolites in the leaves of these 11 accessions were analyzed using LC-MS based untargeted metabolomics. The choice assays showed significant differences among the accessions in thrips feeding damage. The level of resistance depended on plant developmental stage. Metabolomics analyses showed differences in metabolomes among the Capsicum species and plant developmental stages. Moreover, metabolomic profiles of resistant and susceptible accessions differed. Monomer and dimer acyclic diterpene glycosides (capsianosides) were pinpointed as metabolites that were related to thrips resistance. Sucrose and malonylated flavone glycosides were related to susceptibility. To our knowledge, this is the first time that dimer capsianosides of pepper have been linked to insect resistance. Our results show the potential of untargeted metabolomics as a tool for discovering metabolites that are important in plant - insect interactions.


Assuntos
Capsicum/química , Diterpenos/química , Glicosídeos/química , Metabolômica , Animais , Capsicum/metabolismo , Capsicum/parasitologia , Cromatografia Líquida de Alta Pressão , Análise por Conglomerados , Dimerização , Análise Discriminante , Glicosídeos/farmacologia , Interações Hospedeiro-Parasita , Metaboloma , Folhas de Planta/química , Folhas de Planta/metabolismo , Folhas de Planta/parasitologia , Análise de Componente Principal , Espectrometria de Massas em Tandem , Tisanópteros/efeitos dos fármacos , Tisanópteros/fisiologia
20.
J Phys Chem A ; 123(26): 5537-5541, 2019 Jul 05.
Artigo em Inglês | MEDLINE | ID: mdl-31150246

RESUMO

The Raman spectra of polycrystalline samples of ( RS)-2-(4-isobutylphenyl)-propionic acid of the common name ibuprofen have been measured in the temperature range 5-300 K. In the low-frequency spectrum of the normal C12H17(COOH) and deuterated C12H17(COOD) species, modes with ∼103 and ∼95 cm-1 wavenumbers were detected, which corresponded to translational vibrations of O-H(D)···O hydrogen bonds of two different tautomers: left L and right R , respectively. At temperatures below 150 K, only the L-tautomer is found, and at T ≥ 150 K, both tautomers are observed. The energy difference Δ E of the ground vibrational state of potential minima for L- and R-tautomers is ∼80 meV for COOH and ∼70 meV for COOD. At T ≥ 150 K, the vibrational frequency of the C═O bond in the COOH moiety exhibits an unusual temperature dependence.


Assuntos
Dimerização , Ibuprofeno/química , Ligações de Hidrogênio , Modelos Moleculares , Análise Espectral Raman , Temperatura Ambiente , Vibração
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