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1.
Proc Natl Acad Sci U S A ; 120(48): e2312848120, 2023 Nov 28.
Artigo em Inglês | MEDLINE | ID: mdl-37983512

RESUMO

The availability of natural protein sequences synergized with generative AI provides new paradigms to engineer enzymes. Although active enzyme variants with numerous mutations have been designed using generative models, their performance often falls short of their wild type counterparts. Additionally, in practical applications, choosing fewer mutations that can rival the efficacy of extensive sequence alterations is usually more advantageous. Pinpointing beneficial single mutations continues to be a formidable task. In this study, using the generative maximum entropy model to analyze Renilla luciferase (RLuc) homologs, and in conjunction with biochemistry experiments, we demonstrated that natural evolutionary information could be used to predictively improve enzyme activity and stability by engineering the active center and protein scaffold, respectively. The success rate to improve either luciferase activity or stability of designed single mutants is ~50%. This finding highlights nature's ingenious approach to evolving proficient enzymes, wherein diverse evolutionary pressures are preferentially applied to distinct regions of the enzyme, ultimately culminating in an overall high performance. We also reveal an evolutionary preference in RLuc toward emitting blue light that holds advantages in terms of water penetration compared to other light spectra. Taken together, our approach facilitates navigation through enzyme sequence space and offers effective strategies for computer-aided rational enzyme engineering.


Assuntos
Luz , Mutação , Luciferases de Renilla/genética , Luciferases de Renilla/metabolismo , Estabilidade Enzimática
2.
Proc Natl Acad Sci U S A ; 119(31): e2207904119, 2022 08 02.
Artigo em Inglês | MEDLINE | ID: mdl-35901204

RESUMO

Laboratory evolution combined with computational enzyme design provides the opportunity to generate novel biocatalysts. Nevertheless, it has been challenging to understand how laboratory evolution optimizes designer enzymes by introducing seemingly random mutations. A typical enzyme optimized with laboratory evolution is the abiological Kemp eliminase, initially designed by grafting active site residues into a natural protein scaffold. Here, we relate the catalytic power of laboratory-evolved Kemp eliminases to the statistical energy ([Formula: see text]) inferred from their natural homologous sequences using the maximum entropy model. The [Formula: see text] of designs generated by directed evolution is correlated with enhanced activity and reduced stability, thus displaying a stability-activity trade-off. In contrast, the [Formula: see text] for mutants in catalytic-active remote regions (in which remote residues are important for catalysis) is strongly anticorrelated with the activity. These findings provide an insight into the role of protein scaffolds in the adaption to new enzymatic functions. It also indicates that the valley in the [Formula: see text] landscape can guide enzyme design for abiological catalysis. Overall, the connection between laboratory and natural evolution contributes to understanding what is optimized in the laboratory and how new enzymatic function emerges in nature, and provides guidance for computational enzyme design.


Assuntos
Evolução Molecular Direcionada , Enzimas , Engenharia de Proteínas , Catálise , Domínio Catalítico , Entropia , Enzimas/metabolismo , Mutação
3.
Proc Natl Acad Sci U S A ; 119(6)2022 02 08.
Artigo em Inglês | MEDLINE | ID: mdl-35115408

RESUMO

A variety of signals, including inflammasome activation, trigger the formation of large transmembrane pores by gasdermin D (GSDMD). There are primarily two functions of the GSDMD pore, to drive lytic cell death, known as pyroptosis, and to permit the release of leaderless interleukin-1 (IL-1) family cytokines, a process that does not require pyroptosis. We are interested in the mechanism by which the GSDMD pore channels IL-1 release from living cells. Recent studies revealed that electrostatic interaction, in addition to cargo size, plays a critical role in GSDMD-dependent protein release. Here, we determined computationally that to enable electrostatic filtering against pro-IL-1ß, acidic lipids in the membrane need to effectively neutralize positive charges in the membrane-facing patches of the GSDMD pore. In addition, we predicted that salt has an attenuating effect on electrostatic filtering and then validated this prediction using a liposome leakage assay. A calibrated electrostatic screening factor is necessary to account for the experimental observations, suggesting that ion distribution within the pore may be different from the bulk solution. Our findings corroborate the electrostatic influence of IL-1 transport exerted by the GSDMD pore and reveal extrinsic factors, including lipid and salt, that affect the electrostatic environment.


Assuntos
Interleucina-1/metabolismo , Proteínas de Ligação a Fosfato/metabolismo , Proteínas Citotóxicas Formadoras de Poros/metabolismo , Animais , Membrana Celular/metabolismo , Humanos , Inflamassomos/metabolismo , Camundongos , Piroptose/fisiologia , Eletricidade Estática
4.
Proc Natl Acad Sci U S A ; 119(7)2022 02 15.
Artigo em Inglês | MEDLINE | ID: mdl-35135886

RESUMO

Although computational enzyme design is of great importance, the advances utilizing physics-based approaches have been slow, and further progress is urgently needed. One promising direction is using machine learning, but such strategies have not been established as effective tools for predicting the catalytic power of enzymes. Here, we show that the statistical energy inferred from homologous sequences with the maximum entropy (MaxEnt) principle significantly correlates with enzyme catalysis and stability at the active site region and the more distant region, respectively. This finding decodes enzyme architecture and offers a connection between enzyme evolution and the physical chemistry of enzyme catalysis, and it deepens our understanding of the stability-activity trade-off hypothesis for enzymes. Overall, the strong correlations found here provide a powerful way of guiding enzyme design.

5.
J Am Chem Soc ; 146(20): 13875-13885, 2024 May 22.
Artigo em Inglês | MEDLINE | ID: mdl-38718165

RESUMO

Bioluminescence is a fascinating natural phenomenon, wherein organisms produce light through specific biochemical reactions. Among these organisms, Renilla luciferase (RLuc) derived from the sea pansy Renilla reniformis is notable for its blue light emission and has potential applications in bioluminescent tagging. Our study focuses on RLuc8, a variant of RLuc with eight amino acid substitutions. Recent studies have shown that the luminescent emitter coelenteramide can adopt multiple protonation states, which may be influenced by nearby residues at the enzyme's active site, demonstrating a complex interplay between protein structure and bioluminescence. Herein, using the quantum mechanical consistent force field method and the semimacroscopic protein dipole-Langevin dipole method with linear response approximation, we show that the phenolate state of coelenteramide in RLuc8 is the primary light-emitting species in agreement with experimental results. Our calculations also suggest that the proton transfer (PT) from neutral coelenteramide to Asp162 plays a crucial role in the bioluminescence process. Additionally, we reproduced the observed emission maximum for the amide anion in RLuc8-D120A and the pyrazine anion in the presence of a Na+ counterion in RLuc8-D162A, suggesting that these are the primary emitters. Furthermore, our calculations on the neutral emitter in the engineered AncFT-D160A enzyme, structurally akin to RLuc8-D162A but with a considerably blue-shifted emission peak, aligned with the observed data, possibly explaining the variance in emission peaks. Overall, this study demonstrates an effective approach to investigate chromophores' bimolecular states while incorporating the PT process in emission spectra calculations, contributing valuable insights for future studies of PT in photoproteins.


Assuntos
Pirazinas , Teoria Quântica , Pirazinas/química , Pirazinas/metabolismo , Renilla/enzimologia , Luciferases/química , Luciferases/metabolismo , Luminescência , Animais , Imidazóis/química , Benzenoacetamidas
6.
Nature ; 557(7707): 701-705, 2018 05.
Artigo em Inglês | MEDLINE | ID: mdl-29760468

RESUMO

Ion hydration and transport at interfaces are relevant to a wide range of applied fields and natural processes1-5. Interfacial effects are particularly profound in confined geometries such as nanometre-sized channels6-8, where the mechanisms of ion transport in bulk solutions may not apply9,10. To correlate atomic structure with the transport properties of hydrated ions, both the interfacial inhomogeneity and the complex competing interactions among ions, water and surfaces require detailed molecular-level characterization. Here we constructed individual sodium ion (Na+) hydrates on a NaCl(001) surface by progressively attaching single water molecules (one to five) to the Na+ ion using a combined scanning tunnelling microscopy and noncontact atomic force microscopy system. We found that the Na+ ion hydrated with three water molecules diffuses orders of magnitude more quickly than other ion hydrates. Ab initio calculations revealed that such high ion mobility arises from the existence of a metastable state, in which the three water molecules around the Na+ ion can rotate collectively with a rather small energy barrier. This scenario would apply even at room temperature according to our classical molecular dynamics simulations. Our work suggests that anomalously high diffusion rates for specific hydration numbers of ions are generally determined by the degree of symmetry match between the hydrates and the surface lattice.

7.
Nature ; 563(7729): E18, 2018 11.
Artigo em Inglês | MEDLINE | ID: mdl-30135587

RESUMO

In this Letter, the links to Supplementary Videos 5, 7, 9 and 10 were incorrect, and there were some formatting errors in the Supplementary Video legends. These errors have been corrected online.

8.
Sheng Li Xue Bao ; 75(6): 799-816, 2023 Dec 25.
Artigo em Zh | MEDLINE | ID: mdl-38151345

RESUMO

Mitochondria-associated endoplasmic reticulum membranes (MAMs) are the physical connection sites between mitochondria and endoplasmic reticulum (ER). As the compartments controlling substance and information communications between ER and mitochondria, MAMs were involved in the regulation of various pathophysiological processes, such as calcium homeostasis, mitochondrial morphology and function, lipid metabolism and autophagy. In the past decades, accumulating lines of evidence have revealed the pivotal role of MAMs in diverse cardiovascular diseases (CVD). Aging is one of the major independent risk factors for CVD, which causes progressive degeneration of the cardiovascular system, leading to increased morbidity and mortality of CVD. This review aims to summarize the research progress of MAMs in age-related CVD, and explore new targets for its prevention and treatment.


Assuntos
Doenças Cardiovasculares , Membranas Mitocondriais , Humanos , Doenças Cardiovasculares/metabolismo , Sinalização do Cálcio/fisiologia , Mitocôndrias/fisiologia , Retículo Endoplasmático/metabolismo
9.
J Proteome Res ; 21(3): 822-832, 2022 03 04.
Artigo em Inglês | MEDLINE | ID: mdl-34319108

RESUMO

Infantile hemangioma (IH), the most common benign tumor in infancy, mostly arises and has rapid growth before 3 months of age. Because irreversible skin changes occur in the early proliferative stage, early medical treatment is essential to reduce the permanent sequelae caused by IH. Yet there are still no early screening biomarkers for IH before its visible emergence. This study aimed to explore prediction biomarkers using noninvasive umbilical cord blood (UCB). A prospective study of the metabolic profiling approach was performed on UCB sera from 28 infants with IH and 132 matched healthy controls from a UCB population comprising over 1500 infants (PeptideAtlas: PASS01675) using liquid chromatography-mass spectrometry. The metabolic profiling results exhibited the characteristic metabolic aberrance of IH. Machine learning suggested a panel of biomarkers to predict the occurrence of IH, with the area under curve (AUC) values in the receiver operating characteristic analysis all >0.943. Phenylacetic acid had potential to predict infants with large IH (diameter >2 cm) from those with small IH (diameter <2 cm), with an AUC of 0.756. The novel biomarkers in noninvasive UCB sera for predicting IH before its emergence might lead to a revolutionary clinical utility.


Assuntos
Sangue Fetal , Hemangioma , Biomarcadores , Cromatografia Líquida , Hemangioma/complicações , Hemangioma/diagnóstico , Hemangioma/tratamento farmacológico , Humanos , Lactente , Estudos Prospectivos
10.
J Am Chem Soc ; 144(36): 16638-16646, 2022 09 14.
Artigo em Inglês | MEDLINE | ID: mdl-36044733

RESUMO

A variety of diseases are associated with tyrosine kinase enzymes that activate many proteins via signal transduction cascades. The similar ATP-binding pockets of these tyrosine kinases make it extremely difficult to design selective covalent inhibitors. The present study explores the contribution of the chemical reaction steps to the selectivity of the commercialized inhibitor acalabrutinib over the Bruton's tyrosine kinase (BTK) and the interleukin-2-inducible T-cell kinase (ITK). Ab initio and empirical valence bond (EVB) simulations of the two kinases indicate that the most favorable reaction path involves a water-assisted mechanism of the 2-butynamide reactive group of acalabrutinib. BTK reacts with acalabrutinib with a substantially lower barrier than ITK, according to our calculated free-energy profile and kinetic simulations. Such a difference is due to the microenvironment of the active site, as further supported by a sequence-based analysis of specificity determinants for several commercialized inhibitors. Our study involves a new approach of simulating directly the IC50 and inactivation efficiency keff, instead of using the standard formulas. This new strategy is particularly important in studies of covalent inhibitors with a very exothermic bonding step. Overall, our results demonstrate the importance of understanding the chemical reaction steps in designing selective covalent inhibitors for tyrosine kinases.


Assuntos
Benzamidas , Inibidores de Proteínas Quinases , Tirosina Quinase da Agamaglobulinemia , Benzamidas/farmacologia , Inibidores de Proteínas Quinases/química , Inibidores de Proteínas Quinases/farmacologia , Pirazinas , Tirosina
11.
Proc Natl Acad Sci U S A ; 116(6): 2052-2057, 2019 02 05.
Artigo em Inglês | MEDLINE | ID: mdl-30674671

RESUMO

Inflammasomes are large protein complexes that trigger host defense in cells by activating inflammatory caspases for cytokine maturation and pyroptosis. NLRP6 is a sensor protein in the nucleotide-binding domain (NBD) and leucine-rich repeat (LRR)-containing (NLR) inflammasome family that has been shown to play multiple roles in regulating inflammation and host defenses. Despite the significance of the NLRP6 inflammasome, little is known about the molecular mechanism behind its assembly and activation. Here we present cryo-EM and crystal structures of NLRP6 pyrin domain (PYD). We show that NLRP6 PYD alone is able to self-assemble into filamentous structures accompanied by large conformational changes and can recruit the ASC adaptor using PYD-PYD interactions. Using molecular dynamics simulations, we identify the surface that the NLRP6 PYD filament uses to recruit ASC PYD. We further find that full-length NLRP6 assembles in a concentration-dependent manner into wider filaments with a PYD core surrounded by the NBD and the LRR domain. These findings provide a structural understanding of inflammasome assembly by NLRP6 and other members of the NLR family.


Assuntos
Inflamassomos/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Sequência de Aminoácidos , Microscopia Crioeletrônica , Cristalografia por Raios X , Humanos , Imunidade Inata , Peptídeos e Proteínas de Sinalização Intracelular/química , Peptídeos e Proteínas de Sinalização Intracelular/genética , Modelos Moleculares , Simulação de Dinâmica Molecular , Conformação Proteica , Multimerização Proteica
12.
J Minim Access Surg ; 18(4): 585-590, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-36204938

RESUMO

Background: Over the last two decades, several endoscopic thyroidectomy methods have been developed. However, there are some limitations in these procedures. To date, the optimal surgical approach for thyroid cancer has not yet been developed. This study reported the surgical operation steps, clinical outcomes, and experience of 30 patients who underwent trans-cervico-mental angle single-port endoscopic thyroidectomy (TCMASPET) at our centre. Patients and Methods: A total of 30 patients were enrolled in the present study. Patients underwent unilateral or bilateral thyroidectomy through a cervico-mental angle incision of 2.48 ± 0.31 cm, after which the lymphoid adipose tissues in the central region were dissected. Results: All surgeries were successfully completed. Two patients underwent bilateral thyroid carcinoma resection with bilateral central region lymph node dissection, 23 patients received unilateral thyroid cancer resection with unilateral central region lymph node dissection, four patients underwent unilateral thyroid resection, and one patient received bilateral thyroid resection with unilateral central region lymph node dissection. No permanent post-operative complications were observed. Conclusions: TCMASPET was a safe and feasible approach that was relatively easy to perform. This approach may expand the indications for endoscopic thyroidectomy while maintaining excellent cosmetic outcomes.

13.
Angew Chem Int Ed Engl ; 61(44): e202205301, 2022 Nov 02.
Artigo em Inglês | MEDLINE | ID: mdl-35866885

RESUMO

The electrocatalytic CO2 reduction reaction (ECO2 RR) is one promising method for storing intermittent clean energy in chemical bonds and producing fuels. Among various kinds of catalysts for ECO2 RR, molecular metal complexes with well-defined structures are convenient for studies of their rational design, structure-reactivity relationships, and mechanisms. In this Review, we summarize the molecular engineering of several N-based metal complexes including Re/Mn bipyridine compounds and metal macrocycles, concluding with general modification strategies to devise novel molecular catalysts with high intrinsic activity. Through physical adsorption, covalent linking, and formation of a periodic backbone, these active molecules can be heterogenized into immobilized catalysts with more practical prospects. Finally, significant challenges and opportunities based on molecular catalysts are discussed.

14.
PLoS Comput Biol ; 16(9): e1008262, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32986691

RESUMO

Genome organization is critical for setting up the spatial environment of gene transcription, and substantial progress has been made towards its high-resolution characterization. The underlying molecular mechanism for its establishment is much less understood. We applied a deep-learning approach, variational autoencoder (VAE), to analyze the fluctuation and heterogeneity of chromatin structures revealed by single-cell imaging and to identify a reaction coordinate for chromatin folding. This coordinate connects the seemingly random structures observed in individual cohesin-depleted cells as intermediate states along a folding pathway that leads to the formation of topologically associating domains (TAD). We showed that folding into wild-type-like structures remain energetically favorable in cohesin-depleted cells, potentially as a result of the phase separation between the two chromatin segments with active and repressive histone marks. The energetic stabilization, however, is not strong enough to overcome the entropic penalty, leading to the formation of only partially folded structures and the disappearance of TADs from contact maps upon averaging. Our study suggests that machine learning techniques, when combined with rigorous statistical mechanical analysis, are powerful tools for analyzing structural ensembles of chromatin.


Assuntos
Cromatina/metabolismo , Aprendizado Profundo , Cromatina/química , Simulação por Computador , Genoma , Código das Histonas , Modelos Biológicos , Conformação Proteica , Termodinâmica
15.
J Allergy Clin Immunol ; 145(1): 391-401.e8, 2020 01.
Artigo em Inglês | MEDLINE | ID: mdl-31629014

RESUMO

BACKGROUND: Mutations affecting DNA polymerases have been implicated in genomic instability and cancer development, but the mechanisms by which they can affect the immune system remain largely unexplored. OBJECTIVE: We sought to establish the role of DNA polymerase δ1 catalytic subunit (POLD1) as the cause of a primary immunodeficiency in an extended kindred. METHODS: We performed whole-exome and targeted gene sequencing, lymphocyte characterization, molecular and functional analyses of the DNA polymerase δ (Polδ) complex, and T- and B-cell antigen receptor repertoire analysis. RESULTS: We identified a missense mutation (c. 3178C>T; p.R1060C) in POLD1 in 3 related subjects who presented with recurrent, especially herpetic, infections and T-cell lymphopenia with impaired T-cell but not B-cell proliferation. The mutation destabilizes the Polδ complex, leading to ineffective recruitment of replication factor C to initiate DNA replication. Molecular dynamics simulation revealed that the R1060C mutation disrupts the intramolecular interaction between the POLD1 CysB motif and the catalytic domain and also between POLD1 and the Polδ subunit POLD2. The patients exhibited decreased numbers of naive CD4 and especially CD8 T cells in favor of effector memory subpopulations. This skewing was associated with oligoclonality and restricted T-cell receptor ß-chain V-J pairing in CD8+ but not CD4+ T cells, suggesting that POLD1R1060C differentially affects peripheral CD8+ T-cell expansion and possibly thymic selection. CONCLUSION: These results identify gene defects in POLD1 as a novel cause of T-cell immunodeficiency.


Assuntos
DNA Polimerase III , Mutação com Perda de Função , Mutação de Sentido Incorreto , Imunodeficiência Combinada Severa , Adolescente , Motivos de Aminoácidos , Pré-Escolar , DNA Polimerase III/genética , DNA Polimerase III/imunologia , Feminino , Células HEK293 , Humanos , Domínios Proteicos , Imunodeficiência Combinada Severa/genética , Imunodeficiência Combinada Severa/imunologia , Imunodeficiência Combinada Severa/patologia , Sequenciamento do Exoma
16.
Biophys J ; 116(10): 2047-2056, 2019 05 21.
Artigo em Inglês | MEDLINE | ID: mdl-31053260

RESUMO

Epigenetic modifications can extend over long genomic regions to form domain-level chromatin states that play critical roles in gene regulation. The molecular mechanism for the establishment and maintenance of these states is not fully understood and remains challenging to study with existing experimental techniques. Here, we took a data-driven approach and parameterized an information-theoretic model to infer the formation mechanism of domain-level chromatin states from genome-wide epigenetic modification profiles. This model reproduces statistical correlations among histone modifications and identifies well-known states. Importantly, it predicts drastically different mechanisms and kinetic pathways for the formation of euchromatin and heterochromatin. In particular, long, strong enhancer and promoter states grow gradually from short but stable regulatory elements via a multistep process. On the other hand, the formation of heterochromatin states is highly cooperative, and no intermediate states are found along the transition path. This cooperativity can arise from a chromatin looping-mediated spreading of histone methylation mark and supports collapsed, globular three-dimensional conformations rather than regular fibril structures for heterochromatin. We further validated these predictions using changes of epigenetic profiles along cell differentiation. Our study demonstrates that information-theoretic models can go beyond statistical analysis to derive insightful kinetic information that is otherwise difficult to access.


Assuntos
Cromatina/genética , Epigenômica , Modelos Genéticos
17.
J Chem Phys ; 150(12): 124703, 2019 Mar 28.
Artigo em Inglês | MEDLINE | ID: mdl-30927866

RESUMO

We study, in this paper, the physical properties of water confined between two parallel graphene plates with different slit widths to understand the effects of confinement on the water structure and how bulk properties are reached as the water layer thickens. It was found that the microscopic structures of the interfacial liquid layer close to graphene vary with the slit width. Water tends to locate at the center of the six-membered ring of graphene planes to form triangular patterns, as found by others. The narrower the slit width is, the more pronounced this pattern is, except for the slit width of 9.5 Å, for which a well-defined two-layer structure of water forms. On the other hand, squared structures can be clearly seen in single snapshots at small (6.5 Å and 7.5 Å) but not large slit widths. Even at small slit widths, the square-like geometry is observed only when an average is taken for a short trajectory, and averaging over a long time yields a triangular pattern dictated by the graphene geometry. We estimate the length of time needed to observe two patterns, respectively. We also used the two-phase thermodynamic model to study the variation of entropy of confined water and found that at 8.5 Å, the entropy of confined water is larger than that of bulk water. The rotational entropy of confined water is higher than that of bulk water for all slit widths due to the reduction of the hydrogen bond in the confined space.

18.
Biophys J ; 113(7): 1395-1404, 2017 Oct 03.
Artigo em Inglês | MEDLINE | ID: mdl-28978434

RESUMO

The relationship between DNA methylation and chromatin structure is still largely unknown. By analyzing a large set of published sequencing data, we observed a long-range power law correlation of DNA methylation with cell class-specific scaling exponents in the range of tens of kilobases. We showed that such cell class-specific scaling exponents are caused by different patchiness of DNA methylation in different cells. By modeling the chromatin structure using high-resolution chromosome conformation capture data and mapping the methylation level onto the modeled structure, we demonstrated that the patchiness of DNA methylation is related to chromatin structure. The scaling exponents of the power law correlation are thus a display of the spatial organization of chromatin. Besides the long-range correlation, we also showed that the local correlation of DNA methylation is associated with nucleosome positioning. The local correlation of partially methylated domains is different from that of nonpartially methylated domains, suggesting that their chromatin structures differ at the scale of several hundred base pairs (covering a few nucleosomes). Our study provides a novel, to our knowledge, view of the spatial organization of chromatin structure from a perspective of DNA methylation, in which both long-range and local correlations of DNA methylation along the genome reflect the spatial organization of chromatin.


Assuntos
Cromatina/metabolismo , Metilação de DNA/fisiologia , Glândulas Suprarrenais/metabolismo , Animais , Encéfalo/metabolismo , Análise por Conglomerados , Células-Tronco Embrionárias/metabolismo , Feminino , Análise de Fourier , Perfilação da Expressão Gênica , Humanos , Camundongos , Modelos Genéticos , Modelos Moleculares , Neoplasias/genética , Neoplasias/metabolismo , Ovário/metabolismo , Pâncreas/metabolismo
19.
Tumour Biol ; 37(1): 653-9, 2016 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-26242265

RESUMO

Autophagy is an intracellular degradation pathway that delivers organelles or protein to the lysosome and has been recently implicated in the resistance of gastric cancer to chemotherapy. This study aimed to investigate whether blocking autophagy is a new approach for the treatment of chemoresistant gastric cancer. SGC-7901 gastric cancer cell line was treated with 5-fluorouracil (5-FU) or/and autophagy inhibitor bafilomycin A1. Cell viability and growth were evaluated by 3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyl tetrazolium bromide (MTT) and clonogenic assay. Apoptosis was evaluated by flow cytometry. Cell migratory and invasive ability were evaluated by migration and invasion assays. Autophagy was evaluated by scanning electron microscopic, acridine orange staining, and Western blot analysis. We observed that 5-FU induced autophagy in SGC-7901 cells. Bafilomycin A1 decreased the viability and clone formation, inhibited the invasive and migratory ability, and increased the apoptosis of SGC-7901 cells. Taken together, our data suggest that chemotherapy-induced autophagy contributes to gastric cancer chemoresistance, and the inhibition of autophagy is a promising strategy for gastric cancer therapy.


Assuntos
Antineoplásicos/farmacologia , Autofagia/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos , Inibidores Enzimáticos/farmacologia , Macrolídeos/farmacologia , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Proliferação de Células , Sobrevivência Celular , Fluoruracila/farmacologia , Humanos , Neoplasias Gástricas
20.
J Chem Phys ; 143(22): 224504, 2015 Dec 14.
Artigo em Inglês | MEDLINE | ID: mdl-26671387

RESUMO

In this study, we examine how complex ions such as oxyanions influence the dynamic properties of water and whether differences exist between simple halide anions and oxyanions. Nitrate anion is taken as an example to investigate the hydration properties of oxyanions. Reorientation relaxation of its hydration water can occur through two different routes: water can either break its hydrogen bond with the nitrate to form one with another water or switch between two oxygen atoms of the same nitrate. The latter molecular mechanism increases the residence time of oxyanion's hydration water and thus nitrate anion slows down the translational motion of neighbouring water. But it is also a "structure breaker" in that it accelerates the reorientation relaxation of hydration water. Such a result illustrates that differences do exist between the hydration of oxyanions and simple halide anions as a result of different molecular geometries. Furthermore, the rotation of the nitrate solute is coupled with the hydrogen bond rearrangement of its hydration water. The nitrate anion can either tilt along the axis perpendicularly to the plane or rotate in the plane. We find that the two reorientation relaxation routes of the hydration water lead to different relaxation dynamics in each of the two above movements of the nitrate solute. The current study suggests that molecular geometry could play an important role in solute hydration and dynamics.

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