RESUMO
BACKGROUND: Autoimmune disorders encompass a diverse subset of diseases whose common symptoms include, among others, fever. Fever of unknown origins, once an infectious or tumor agent have been ruled out as possible causes, may originate with an autoimmune disease. OBJECTIVE: To determine the role of febrile temperatures on the stability of antigens pertinent to autoimmunity, and on the immune complexes they form with commercial therapeutic monoclonal antibodies. METHODS: Using molecular dynamics simulations, the binding between four antigens belonging to a set of autoimmune diseases and their individual monoclonal antibodies was investigated under different febrile temperatures. RESULTS: It was determined that at febrile temperatures, monoclonal antibodies used in the therapy of autoimmune diseases bind with higher binding free energy to pertinent antigens, once the autoimmune condition has been established and treatment is warranted. CONCLUSION: Performing molecular dynamics simulations at fever temperatures may be important for delineating the role antibodies may play in other diseases, including in cancers and infections.
Assuntos
Complexo Antígeno-Anticorpo , Doenças Autoimunes , Humanos , Temperatura , Autoimunidade , Antígenos , Febre , Anticorpos MonoclonaisRESUMO
The ability to memorize changes in the environment is present at all biological levels, from social groups and individuals, down to single cells. Trans-generational memory is embedded subcellularly through genetic and epigenetic mechanisms. Evidence that cells process and remember features of the immediate environment using protein sensors is reviewed. It is argued that this mnemonic ability is encapsulated within the protein conformational space and lasts throughout its lifetime, which can overlap with the lifespan of the organism. Means to determine diachronic changes in protein activity are presented.
Assuntos
Complexos Multiproteicos/metabolismo , Proteínas/química , Adaptação Biológica , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Proteína Quinase Tipo 2 Dependente de Cálcio-Calmodulina/química , Proteína Quinase Tipo 2 Dependente de Cálcio-Calmodulina/metabolismo , Meia-Vida , Ligantes , Complexos Multiproteicos/química , Conformação Proteica , Proteínas/metabolismo , Transdução de SinaisRESUMO
Protein-folding stress is characteristic of specialized secretory cells and plays a dominant role in a multitude of diseases. The unfolded protein response (UPR) thus triggered is a proteostatic signaling network that adapts the protein-folding capacity of the endoplasmic reticulum to the cellular demands. We have measured the binding affinities between human GRP78, an essential chaperone located in ER, and two transmembrane UPR sensors (human PERK and Ire1α), with or without the addition of an unfolded protein client. We reveal distinct binding affinities between the binary and ternary complexes thus formed, that suggest a preference for the PERK signaling branch under stress, and a predilection for the GRP78-UPR sensor complex formation upon stressor removal. These results imply a gated UPR mechanism that tunes the overall cellular behavior to the accumulation of unfolded proteins.
Assuntos
Endorribonucleases/química , Proteínas de Choque Térmico/química , Simulação de Acoplamento Molecular , Proteínas Serina-Treonina Quinases/química , Resposta a Proteínas não Dobradas , eIF-2 Quinase/química , Sítios de Ligação , Chaperona BiP do Retículo Endoplasmático , Endorribonucleases/ultraestrutura , Proteínas de Choque Térmico/ultraestrutura , Humanos , Modelos Químicos , Ligação Proteica , Proteínas Serina-Treonina Quinases/ultraestrutura , eIF-2 Quinase/ultraestruturaRESUMO
The self-assembly and electron transfer properties of adsorbed organic molecules are of interest for the construction of miniaturized molecular circuitries. We have investigated with scanning probe microscopy the self-organization of two structurally related molecular wires embedded within a supportive alkanethiol matrix. Our results evidence heterogeneous adsorption patterns of the molecular wires on gold with either incommensurate unit cells driven into assembly by lateral interactions or a dynamic, commensurate distribution on gold, along with formation of distinct 2D phases. We also observed diffusion-based conductance switching for one of the molecular wires, due to its propensity toward weaker lateral interactions and Au-S adatom formation. We have further demonstrated through the use of scanning tunneling spectroscopy differential current-voltage response for each molecular wire, despite their close structural similarity. Such molecular wires embedded in alkanethiol matrix and exhibiting conductance-switching phenomena have the potential to be used for the functionalization of electrodes in bioelectronic devices.
RESUMO
BACKGROUND: Symptoms of rheumatoid arthritis are associated with local inflammation and may include low-grade fever. OBJECTIVE: To establish the role of fever/inflammation temperatures (38â-39â) on the activity of autoantibodies and therapeutic antibodies relevant for rheumatoid arthritis. METHODS: Through the use of molecular dynamics and free energy calculations, we investigated the role of temperature on the formation of pertinent immune complexes. RESULTS: Rheumatoid arthritis autoantibodies bind with higher affinity at febrile/inflammation temperatures. CONCLUSIONS: Fever may modulate binding affinity of autoantibodies relevant for rheumatoid arthritis.
Assuntos
Artrite Reumatoide , Autoanticorpos , Humanos , Temperatura , InflamaçãoRESUMO
Progression of various cancers and autoimmune diseases is associated with changes in systemic or local tissue temperatures, which may impact current therapies. The role of fever and acute inflammation-range temperatures on the stability and activity of antibodies relevant for cancers and autoimmunity is unknown. To produce molecular dynamics (MD) trajectories of immune complexes at relevant temperatures, we used the Research Collaboratory for Structural Bioinformatics (RCSB) database to identify 50 antibody:antigen complexes of interest, in addition to single antibodies and antigens, and deployed Groningen Machine for Chemical Simulations (GROMACS) to prepare and run the structures at different temperatures for 100-500 ns, in single or multiple random seeds. MD trajectories are freely available. Processed data include Protein Data Bank outputs for all files obtained every 50 ns, and free binding energy calculations for some of the immune complexes. Protocols for using the data are also available. Individual datasets contain unique DOIs. We created a web interface, ThermoPCD, as a platform to explore the data. The outputs of ThermoPCD allow the users to relate thermally-dependent changes in epitopes:paratopes interfaces to their free binding energies, or against own experimentally derived binding affinities. ThermoPCD is a free to use database of immune complexes' trajectories at different temperatures that does not require registration and allows for all the data to be available for download. Database URL: https://sites.google.com/view/thermopcd/home.
Assuntos
Simulação de Dinâmica Molecular , Neoplasias , Humanos , Complexo Antígeno-Anticorpo/química , Temperatura , Sítios de Ligação de AnticorposRESUMO
HER2 and HER3 receptors dimerize into potent pro-oncogenic complexes involved in various aggressive and recurrent tumors. The role of febrile temperatures on the formation of HER2:HER3 complexes is unknown. To this end, molecular dynamics simulations of HER2 and HER3 were performed in the 37 °C-40 °C range. HER2 and ligand-free HER32 display inactive conformers that cannot form complexes at 40 °C, while maintaining their extended conformations able to dimerize in the 37 °C-39 °C range. Thermal therapy at particular fever points may complement existing therapy options for HER2-relevant cancers.Communicated by Ramaswamy H. Sarma.
RESUMO
Aim: Fever follows the administration of trastuzumab and pertuzumab used in HER2-relevant immunotherapy, but is often eliminated in clinical practice. This work explores the role of temperature (37-39°C) in the formation of immune complexes between HER2 with either trastuzumab or pertuzumab or with both antibodies. Materials & methods: Using molecular dynamics simulations and free energy calculations, the binding between HER2 and these immunotherapeutic monoclonal antibodies was investigated at different temperatures. Results: Trastuzumab and pertuzumab present the highest binding free energy to HER2 at febrile temperatures (39°C), or when HER2 is in complex with both antibodies. Conclusion: Performing molecular dynamics simulations under fever temperatures may be important for delineating their role in enhancing the binding affinity of mature antibodies used in immunotherapy.
Breast cancer patients may present fever due to the cancer itself, due to treatment with chemotherapy or monoclonal antibody therapy, or after surgery. In this work, the role of febrile temperatures on the activity of two of the most commonly used monoclonal antibodies for breast cancer treatment, trastuzumab and pertuzumab, was investigated. These therapeutic agents benefit from fever in terms of binding to their tumor target, particularly when both antibodies are deployed together, mirroring the clinical benefits of their dual therapy. These results are important because, in clinical practice, fever that accompanies treatment in cancer patients is usually eliminated. As such, further investigations into the positive role of fever-range temperatures in assisting antibody therapy for breast cancer are warranted.
Assuntos
Neoplasias da Mama , Receptor ErbB-2 , Humanos , Feminino , Trastuzumab/uso terapêutico , Temperatura , Anticorpos Monoclonais Humanizados/uso terapêutico , Neoplasias da Mama/tratamento farmacológico , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêuticoRESUMO
Using the bright, photostable, charged and hydrophilic fluorescent dye Alexa 488 hydrazide to label the fluid phase around intact guard cells, we show that these cells incorporate the fluid phase during constitutive endocytosis against the high turgor. Mobile, cortical and diffraction-limited signals were not observed if a concentration <4 mm was used to stain the fluid phase, suggesting that endocytic vesicles had to be loaded with a minimal number of dye molecules to produce a signal above the background. To quantify the number of molecules taken up by the vesicles, we prepared liposomes, filled with various concentrations of Alexa 488 hydrazide, fractionated them according to their size and imaged them under identical conditions as the guard cells. From the size/intensity relations of these liposomes, we extrapolated the molecular brightness of Alexa 488 hydrazide. Using this calibration, the mean fluorescent intensity of single endocytic vesicles translates into a mean number of 573 Alexa 488 molecules. If a vesicle needs to take up 573 molecules from a 4 mm solution, it requires a diameter of at least 87 nm. This number provides the first in vivo estimate for the size of endocytic vesicles in intact, turgid plant cells.
Assuntos
Endocitose , Corantes Fluorescentes/análise , Hidrazinas/análise , Vesículas Transportadoras/fisiologia , Células Cultivadas , Lipossomos/administração & dosagem , Microscopia de Fluorescência/métodos , Coloração e Rotulagem/métodos , Vicia faba/fisiologiaRESUMO
Hemolysis modulates susceptibility to bacterial infections and predicts poor sepsis outcome. Hemolytic bacteria use hemolysins to induce erythrocyte lysis and obtain the heme that is essential for bacterial growth. Hemolysins are however potent immunogens and infections with hemolytic bacteria may cause a reversible fever response from the host that will aid in pathogen clearance. We hypothesized that fever temperatures impact the growth and infectivity of two hemolytic bacteria that are known to evoke fever in patients. To that end, we used high-sensitivity microcalorimetry to measure the evolution of heat production in fever-inducing strains of Escherichia coli and Staphylococcus aureus, under different temperature conditions. We determined specific bacterial aggregation profiles at temperatures equal to or exceeding 38.5 °C. Two melting temperatures peaks ranged from 38 °C to 43 °C for either species, a feature that we assigned to the formation of hemolysin aggregates of different oligomerization order. In order to measure the role of fever temperatures on hemolysis, we incubated the pathogens on blood agar plates at relevant temperatures, measuring the presence of hemolysis at 37 °C and its absence at 40.5 °C, respectively. We conclude that fever temperatures affect the kinetics of hemolysin pore formation and subsequently the hemolysis of red blood cells in vitro. We reveal the potential of microcalorimetry to monitor heat response from fever inducing bacterial species. Furthermore, these results help establish an additional positive role of febrile temperatures in modulating the immune response to infections, through the abolishment of hemolysis.
RESUMO
Fever is a regulated increase of the body temperature resulting from both infectious and non-infectious causes. Fever is known to play a role in modulating immune responses to infection, but the potential of febrile temperatures in regulating antigen binding affinity to antibodies has not been explored. Here we investigated this process under in vitro conditions using Isothermal titration calorimetry and ELISA. We used selected malarial and dengue antigens against specific monoclonal antibodies, and observed a marked increase in the affinity of these antibody-antigen complexes at 40°C, compared to physiological (37°C) or pathophysiological temperatures (42°C). Induced thermal equilibration of the protein partners at these temperatures in vitro, prior to measurements, further increased their binding affinity. These results suggest another positive and adaptive role for fever in vivo, and highlight the favourable role of thermal priming in enhancing protein-protein affinity for samples with limited availability.
Assuntos
Anticorpos Antivirais/imunologia , Afinidade de Anticorpos , Antígenos Virais/imunologia , Febre/imunologia , Temperatura , Anticorpos Monoclonais/imunologia , Complexo Antígeno-Anticorpo/imunologia , Temperatura Corporal , Calorimetria , Dengue/imunologia , Vírus da Dengue , Ensaio de Imunoadsorção Enzimática , Interações Hospedeiro-Patógeno , Humanos , Malária/imunologia , Plasmodium vivaxRESUMO
OBJECTIVE: Oscillations of physiological parameters describe many biological processes and their modulation is determinant for various pathologies. In sepsis, toll-like receptor 4 (TLR4) is a key sensor for signaling the presence of Gram-negative bacteria. Its intracellular trafficking rates shift the equilibrium between the pro- and anti-inflammatory downstream signaling cascades, leading to either the physiological resolution of the bacterial stimulation or to sepsis. This study aimed to evaluate the effects of TLR4 increased expression and intracellular trafficking on the course and outcome of sepsis. RESULTS: Using a set of three differential equations, we defined the TLR4 fluxes between relevant cell organelles. We obtained three different regions in the phase space: (1) a limit-cycle describing unstimulated physiological oscillations, (2) a fixed-point attractor resulting from moderate LPS stimulation that is resolved and (3) a double-attractor resulting from sustained LPS stimulation that leads to sepsis. We used this model to describe available hospital data of sepsis patients and we correctly characterize the clinical outcome of these patients.