In this study,we present the characterization of the carbon fibers recovered from the mechanochemical-enhanced recycling of carbon fiber reinforced fibers.The objectives of the study were to investigate the effect of ...In this study,we present the characterization of the carbon fibers recovered from the mechanochemical-enhanced recycling of carbon fiber reinforced fibers.The objectives of the study were to investigate the effect of our modified recycling method on the interfacial properties of recovered fibers.The reinforced plastics were recycled;the recycling efficiency was determined and the recovered fibers were sized using 1 wt%and 3 wt%concentration of(3-aminopropyl)triethoxysilane.We characterized the morphologies utilizing the electron spectroscopy for chemical analysis(ESCA),atomic force microscopy(AFM),FTIR-attenuated total reflection(ATR)spectroscopy and scanning electron microscopy(SEM).Although the surface of the fibers had no cracks,there was evidence of contaminations which affected the interfacial properties and the quality of the fibers.Results showed that the trends in the recovered and virgin fibers were similar with an increase in sizing concentration.The results highlighted the perspectives of increasing the quality of recovered fibers after the recycling process.展开更多
Activated carbon was prepared from bamboo by mechanochemical process with phosphoric acid as activating agent.The effects of milling time on the adsorption property and porous structure of the obtained activated car-b...Activated carbon was prepared from bamboo by mechanochemical process with phosphoric acid as activating agent.The effects of milling time on the adsorption property and porous structure of the obtained activated car-bon were discussed.The results showed that phosphoric acid activation assisted by mechanochemical process can slightly improve the adsorption performance of the prepared activated carbon.The iodine,methylene blue adsorption value and the specific surface area of the prepared activated carbons increased from 840.16 mg/g,168 mg/g and 1229.246 m/g to 947.51 mg/g,195 mg/g,and 1265.373 m2/g,respectively.Compared with conventional phosphoric acid activation,the activated carbon produced by mechanochemical process has greater adsorption capacity.展开更多
Embryonic stem (ES) cell biology is attracting much attention in cell biology because of their pluripotent behaviors and potential therapeutic applications. However,what maintains ES cell pluripotency and what trigger...Embryonic stem (ES) cell biology is attracting much attention in cell biology because of their pluripotent behaviors and potential therapeutic applications. However,what maintains ES cell pluripotency and what triggers ES cell展开更多
A new technology of treating molybdenum residues by simultaneous ultrafine milling and alkali leaching was put forward to recover molybdenum from metallurgical residues. The effects of residue size, milling time, soli...A new technology of treating molybdenum residues by simultaneous ultrafine milling and alkali leaching was put forward to recover molybdenum from metallurgical residues. The effects of residue size, milling time, solid content, n (Na 2CO 3)/ n (Mo) and slurry pH value on molybdenum leaching rate were investigated. The results indicate that a simpler process, lower slurry temperature, 50% shorter treating time, 60% decrease of Na 2CO 3 content and 15% increase of molybdenum leaching rate can be obtained by the new technology compared with the traditional process. The leaching kinetic equation was determined, and calculation of active energy ( E =56.2 kJ/mol) shows that the leaching process of molybdenum residues by simultaneous ultrafine milling and alkali leaching is controlled by chemical reaction. Potential exists for the new process to form the basis for an economically viable, environmentally friendly process to recover valuable elements from residues.展开更多
Von Willebrand Factor(VWF),a multimeric plasma glycoprotein,is synthesized in endothelial cells and megakaryocytes.In adhesion and aggregation of circulating platelets towards to the sites of vascular injury,VWF captu...Von Willebrand Factor(VWF),a multimeric plasma glycoprotein,is synthesized in endothelial cells and megakaryocytes.In adhesion and aggregation of circulating platelets towards to the sites of vascular injury,VWF captures and activates the circulating platelets through interaction with platelet GPlba.As a triplet complex of A1A2A3,the VWF-A domain is a closed conformation with a low affinity to GPlba,but mutations or pathological hemodynamic environment of high fluid shear stress can induce the closed A domain to become an extended one.However,the key events in the force-and/or mutation-induced activation of VWF-A under flows remains unclear.Therefore,with techniques of AFM and PPFC,we here examined transformation of conformation and function of VWF-A under various wall shear stresses,for understanding regulation of force on VWF-A activation.Interesting,AFM scanning imaging data showed that VWF-A molecules on substrate pretreated by perfusing distilled water at various wall shear stresses shortened first and then lengthened as increasing of the pre-loaded wall shear stress,and the threshold of the wall shear stress is about 100 dyn/cm2,demonstrating that increasing pre-loaded wall shear stress would make the treated-A1A2A3 conformation gradually transform from a loose spherical structure to a compact one first and then become an open or extended one.The adhesion frequency of GPlba-coated Polystyrene microspheres(3-μm radius)on the VWF-A-coated substrates decreased first and then increased with the preloaded wall shear stress,which has a same threshold mentioned above.These results suggested that,force-induced activation of VWF-A occurs just at high wall shear stresses(>100 dyn/cm2).The mechanical stability of the closed A1A2A3 conformation would be weakened by the gain of function(GOF)mutant R1 308 L of A1 and enhanced by the loss of function(LOF)mutant G1324S,as it should be.To further reveal the molecular mechanism of the force-induced enhancing or weakening of VWF-A activation,we performed AFM experiment to investigate interaction of A1(WT A1 and its two mutants,the GOF mutant R1 308 L and the LOF mutant G1324S)with A2 and A3,respectively.The adhesive frequency of A1 with A2was larger than that of A1 with A3,showing that A1 was in favor of A2 rather than A3.And,the lifetimes of A1-A2 and A1-A3bond were biphasic force-dependent,showing a'Catch-slip bond'transform in binding of A1 to A2 or A3.It suggested that under the low wall shear stresses,force could inhibit VWF activity through a catch bond mechanism,which enhanced the stability of the closed A1A2A3 conformation,but under high wall shear stresses,the force would enhance VWF activity through a slip bond mechanism,which promoted conformational transform of VWF-A from closed to extended one through reducing the stability of the closed A1A2A3 structure.Our results showed that the GOF mutant R1 308 L would down-regulate the binding affinity of A1 to A2,leading to a low barrier in opening of the closed VWF-A structure.In contrast,the LOF mutant G1324S would enhance the stability of the closed VWF-A conformation by up-regulating the binding affinity of A1 to A2,leading to inhibition of VWF activity.展开更多
The characteristics of morphologies and changes of the microstructures in mechanically alloyed （MA） （m（Cr）∶m（C）∶m（Cr）=90∶1∶9） mixed powders were investigated by using SEM observation and X-ray diffracti...The characteristics of morphologies and changes of the microstructures in mechanically alloyed （MA） （m（Cr）∶m（C）∶m（Cr）=90∶1∶9） mixed powders were investigated by using SEM observation and X-ray diffraction （XRD） analyses. The results show that Cu, C and Cr mixed powders mechanically alloyed for 12 h forms the Cu-based super-saturated solid solution; after high energy ball-milling for 96 h the partial solute elements Cr and C precipitate, meanwhile, the mechanochemical reactions take place and the carbides Cr7C3 form. The composite powders after mechanically alloyed are fine and flat. The formed defects such as free surface, grain boundary, sub-grain boundary and dislocation decrease diffusion activity energy, which is the important reason for developing super-saturated solid solution and promoting the second phase precipitation.展开更多
Integrin activation,the transition from a low to a high affinity state,regulates the numerous cellular responses consequent to integrin engagement by extracellular matrix proteins.Kindlin proteins,play crucial roles i...Integrin activation,the transition from a low to a high affinity state,regulates the numerous cellular responses consequent to integrin engagement by extracellular matrix proteins.Kindlin proteins,play crucial roles in the integrin-signaling pathway by directly interacting with and activating integrins,which mediate the cell-extracellular matrix adhesion and signaling.As a widely distributed PTB domain protein and a major member of the kindlin family,kindlin2 interacts withβ3-tail,bridges talin-activated integrins to promote integrin aggregation,and enhances talin-induced integrin activation.Thus,kindlin2 is identified as a coactivator of integrins.Unlike talins,kindlin2 cannot directly alter the conformation of the integrin transmembrane helix and fail to activate integrin alone.Nevertheless,although it is widely accepted that kindlins and talins synergistically promote integrin activation,the underlying mechanism is unclear.Thus,the study of the force dissociation of the kindlin2/β3-tail complex and the conformation stabilization under different mechanical micro-environments should be of great significance for the further understanding of the structural basis of its synergistically activation of integrin.To reveal the molecular dynamics mechanism of interaction between kindlin2 andβ3-tail,we perform molecular dynamics(MD)simulations for this complex with different computing strategies interaction.In MD simulations,the available crystal structures of Kindlin-2/β3-tail complex(Protein Data Bank code 5XQ1)was downloaded from the PDB database.Two software packages,VMD for visualization and modeling and NAMD 2.13 for energy minimizations and MD simulations,were used here.The steadystate conformation of the complex was obtained from the equilibrium simulation.The dissociation event was observed by the constant velocity simulation,and the mechanical stability of the complex was observed by the constant force simulation.Our results showed that,during the equilibrium of the kindlin2-F3/β34ail complex,the residue MET612,LYS613 and TRP615 on the F3 domain of kindlin2 contributed to hydrogen-bonding with the corresponding residues onβ3 integrin.These bonds exhibit moderate or strong stability through steered molecular dynamics(SMD)simulation.During the constant velocity simulation,the complex exhibits a variety of unfolding pathways against tension applications,which are mainly distinguished by the disruption of hydrogen-bonds between the F3 domain a1/a2 helixes andβ1/β2 sheets.During the constant force simulation,the different phases of the composite force dissociation have different dissociation probabilities,which shows the biphasic force-dependent characteristics.And,the key residues in the pulling were recognized according not only to the number of interacting residue pairs,but also to their bond strength.Using molecular dynamics simulation,we showed the steady state of the kindlin2-F3/β3-tail complex under different tensile forces,and observe the dynamic process of molecular interaction.A possible underlying biophysical mechanism is that,the dissociation of Kindlin2-F3/β3-tail complex is biphasic force-dependent,and the conformations under different stretching states have different binding affinities.This study not only provides insights into the structural basis and mechanical regulation mechanisms of the kindlin/integrin interaction,in understanding in kindlin/integrin-related signaling in different cellular biological processes,but also provides new ideas for novel drug design and the treatment of related diseases.展开更多
CD40 receptor and its ligand CD40 L are the members of TNFR and TNF family,respectively.CD40 is expressed on antigen-presenting cells(APC)as a monomer,and would be dimerized upon its oligomerization through binding to...CD40 receptor and its ligand CD40 L are the members of TNFR and TNF family,respectively.CD40 is expressed on antigen-presenting cells(APC)as a monomer,and would be dimerized upon its oligomerization through binding to its ligand CD40 L.Such dimer formation was shown to be essential for some CD40 signaling events,including phosphoinositide-3 kinase(PI-3 K)activation,the subsequent B7-2 upregulation and the production of IL-8.CD40 L is primarily expressed on activated T cells and activated platelets,and acts as an important costimulatory molecule especially on T follicular helper cells.It is believed that CD40-CD40 L interaction deeply involved in many immune events for the host defense against pathogens and cancer.CD40-CD40 L interaction mediates intracellular signaling along different pathways,such as the canonical and noncanonical nuclear factorκB pathway,mitogen-activated protein kinases,phosphatidylinositol-3 kinase(PI3K)and the phospholipase Cy pathway,is necessary for successful adaptive immune responses mainly relevant to development of CD8+CTLs,and is deeply involved in cross-talks among T cell,B cell,tumor cell,platelet and etc.However,interaction of CD40 with platelet CD40 L would occur in hemodynamics environments,and the regulation of force on this interaction remain unclear.Besides polymerization of CD40,the hemodynamic environment could be believed to regulate interaction between CD40 and its ligand.To reveal the mechanical regulation mechanism of interaction between CD40 L and CD40,atomic force microscope(AFM)was used in the adhesion frequency assay for measuring two-dimensional(2D)kinetics of CD40 L with CD40(the monomer and its dimer)at zero-force and force-clamp assay for measuring single-bond lifetimes of CD40 L/CD40 complex in a range of forces at single-molecule level.AFM cantilever probes were functionalized by incubation with CD40 L in buffer,and soaked in phosphate-buffered saline(PBS)containing 1%bovine serum albumin(BSA)to block nonspecific binding.Polystyrene dishes were coated with CD40(the monomer and its dimer)and then filled with PBS containing 1%BSA.Our data showed that the adhesion frequency of CD40 L with CD40 monomer and its dimer was 0.10 and0. 15,respectively,showing a higher affinity of CD40 L to CD40 dimer rather than its monomer;the rupture force for CD40 L dissociating from CD40 monomer and its dimer was 23 and 31pN,respectively,demonstrating a higher mechanical strength of complex of CD40 L with CD40 dimer instead of monomer;and increasing tension will prolong first and then shorten the bond lifetime of CD40 L-CD40 complex with force threshold of about 1OpN,and dimerization of CD40 had significantly increased enhanced the bond lifetime of CD40 L-CD40 complex,showing a dimerization-enhanced affinity of CD40 to CD40 L and'Catch-Slip'bond mechanism of CD40 L-CD40 interaction.These results suggest that,through upregulating the binding affinity with CD40 L,the polymerization of CD40 stabilizes the linkers and thereby communication between cell and cell,so does the force being small than the force threshold Both CD40 polymerization-and'Catch bond'-induced enhancement of affinity of CD40 with CD40 L may be necessary for stable cross talks between two cells.This finding may be useful for understanding CD40/CD40 L-induced events importantly in physiological or pathological processes at molecular and cellular level.展开更多
As a key regulator of immune response,CD40 L is usually associated with chronic disease-related inflammation,autoimmune diseases and malignant diseases.Receptor recognition of platelet CD40 L is the initial event that...As a key regulator of immune response,CD40 L is usually associated with chronic disease-related inflammation,autoimmune diseases and malignant diseases.Receptor recognition of platelet CD40 L is the initial event that mediates platelet aggregation and leukocyte immune response.Unlike soluble CD40 L,the interaction between transmembrane platelet CD40 L and its receptors occurs within the cell junction surface,usually,in a physiological and pathological high blood flow shear stress environment.This two-dimensional reaction kinetics should be a mechano-chemical coupling process.In addition to its classical receptor CD40,CD40 L also binds to receptorα5β1,CD40 L can bind to the resting state of integrinα5β1,but the mechanical regulation mechanism of integrinα5β1 activation under fluid shear stress remains unclear.We assume that the force can promote CD40 L-inducedα5β1 activation.To check this hypothesis,we performed flow chamber experiment to investigate interaction of CD40 L andα5β1.In experiments,the bottom of the flow chamber is functionalized by a suitable concentration of CD40 L,and the fiber spheres of 6μm diameter was coated withα5β1.The selection of CD40 L concentration was based on the observation that as many tether events ofα5β1-coated spheres as possible were observed rather than stable adhesion events of these spheres.Theα5β1-coated sphere suspension was poured over the CD40 L-coated substrates in the flow chamber under different shear rates.A high-speed camera was used to observe and record tether events of fiber spheres at a rate of 100 frames per second.According to our affinity state transition model for integrin,the data were analyzed to obtain the rate of integrin activation and its mechanical regulation characteristics.Our results demonstrated that the interaction betweenα5β1 and CD40 L is biphasic force-dependent,showing mechano-chemical regulation mechanism of'Catch-slip bond'transition.The affinity jumping model was well fitted with the data obtained from flow chamber experiment at various wall shear stresses.We found that,CD40 L ligation-induced jumping ofα5β1 affinity state from low to medium(or high)one will occur within 0.5-1.0 second,resulting in prolonging of bond lifetimes.And,frequency distribution of the tether events number with tether lifetime under each force,exhibits obvious doublet peaks,one within 0.5-1 s and second within 1.5-2.5 s,indicating theα5β1 affinity state transform from low to high one.The probability distribution of the tether lifetime under different shear forces are not linear,and exists a turning point,which shows that the rate ofα5β1 dissociation from CD40 L is fast first,and then become slow,showing a force-induced conformation transformation of the integrinα5β1 from low affinity state to high affinity one.Our findings suggest that,the continuous force stimulation will quickly cause the affinity state change of integrinα5β1. The dissociation rate of theα5β1/CD40 L complex decreases first and then increases with wall shear stress,exhibiting a'Catch-slip bond'transformation of interaction betweenα5β1-CD40 L.This mechanical regulation mechanism exists in interaction of CD40 L not only toα5β1 at low affinity state but also to one at high affinity state.Our results should be useful in understanding the mechanical regulation mechanism of a5β1-CD40 L interaction-mediated cellular immune response and inflammatory processes.展开更多
基金Project(S2598445)supported by the Project for Cooperative R&D between Industry,Academy and Research Institute Funded by the Korea Ministry of SME and Startups in 2018
文摘In this study,we present the characterization of the carbon fibers recovered from the mechanochemical-enhanced recycling of carbon fiber reinforced fibers.The objectives of the study were to investigate the effect of our modified recycling method on the interfacial properties of recovered fibers.The reinforced plastics were recycled;the recycling efficiency was determined and the recovered fibers were sized using 1 wt%and 3 wt%concentration of(3-aminopropyl)triethoxysilane.We characterized the morphologies utilizing the electron spectroscopy for chemical analysis(ESCA),atomic force microscopy(AFM),FTIR-attenuated total reflection(ATR)spectroscopy and scanning electron microscopy(SEM).Although the surface of the fibers had no cracks,there was evidence of contaminations which affected the interfacial properties and the quality of the fibers.Results showed that the trends in the recovered and virgin fibers were similar with an increase in sizing concentration.The results highlighted the perspectives of increasing the quality of recovered fibers after the recycling process.
文摘Activated carbon was prepared from bamboo by mechanochemical process with phosphoric acid as activating agent.The effects of milling time on the adsorption property and porous structure of the obtained activated car-bon were discussed.The results showed that phosphoric acid activation assisted by mechanochemical process can slightly improve the adsorption performance of the prepared activated carbon.The iodine,methylene blue adsorption value and the specific surface area of the prepared activated carbons increased from 840.16 mg/g,168 mg/g and 1229.246 m/g to 947.51 mg/g,195 mg/g,and 1265.373 m2/g,respectively.Compared with conventional phosphoric acid activation,the activated carbon produced by mechanochemical process has greater adsorption capacity.
文摘Embryonic stem (ES) cell biology is attracting much attention in cell biology because of their pluripotent behaviors and potential therapeutic applications. However,what maintains ES cell pluripotency and what triggers ES cell
文摘A new technology of treating molybdenum residues by simultaneous ultrafine milling and alkali leaching was put forward to recover molybdenum from metallurgical residues. The effects of residue size, milling time, solid content, n (Na 2CO 3)/ n (Mo) and slurry pH value on molybdenum leaching rate were investigated. The results indicate that a simpler process, lower slurry temperature, 50% shorter treating time, 60% decrease of Na 2CO 3 content and 15% increase of molybdenum leaching rate can be obtained by the new technology compared with the traditional process. The leaching kinetic equation was determined, and calculation of active energy ( E =56.2 kJ/mol) shows that the leaching process of molybdenum residues by simultaneous ultrafine milling and alkali leaching is controlled by chemical reaction. Potential exists for the new process to form the basis for an economically viable, environmentally friendly process to recover valuable elements from residues.
基金supported by the National Natural Science Foundation of China ( 116272109,11432006)
文摘Von Willebrand Factor(VWF),a multimeric plasma glycoprotein,is synthesized in endothelial cells and megakaryocytes.In adhesion and aggregation of circulating platelets towards to the sites of vascular injury,VWF captures and activates the circulating platelets through interaction with platelet GPlba.As a triplet complex of A1A2A3,the VWF-A domain is a closed conformation with a low affinity to GPlba,but mutations or pathological hemodynamic environment of high fluid shear stress can induce the closed A domain to become an extended one.However,the key events in the force-and/or mutation-induced activation of VWF-A under flows remains unclear.Therefore,with techniques of AFM and PPFC,we here examined transformation of conformation and function of VWF-A under various wall shear stresses,for understanding regulation of force on VWF-A activation.Interesting,AFM scanning imaging data showed that VWF-A molecules on substrate pretreated by perfusing distilled water at various wall shear stresses shortened first and then lengthened as increasing of the pre-loaded wall shear stress,and the threshold of the wall shear stress is about 100 dyn/cm2,demonstrating that increasing pre-loaded wall shear stress would make the treated-A1A2A3 conformation gradually transform from a loose spherical structure to a compact one first and then become an open or extended one.The adhesion frequency of GPlba-coated Polystyrene microspheres(3-μm radius)on the VWF-A-coated substrates decreased first and then increased with the preloaded wall shear stress,which has a same threshold mentioned above.These results suggested that,force-induced activation of VWF-A occurs just at high wall shear stresses(>100 dyn/cm2).The mechanical stability of the closed A1A2A3 conformation would be weakened by the gain of function(GOF)mutant R1 308 L of A1 and enhanced by the loss of function(LOF)mutant G1324S,as it should be.To further reveal the molecular mechanism of the force-induced enhancing or weakening of VWF-A activation,we performed AFM experiment to investigate interaction of A1(WT A1 and its two mutants,the GOF mutant R1 308 L and the LOF mutant G1324S)with A2 and A3,respectively.The adhesive frequency of A1 with A2was larger than that of A1 with A3,showing that A1 was in favor of A2 rather than A3.And,the lifetimes of A1-A2 and A1-A3bond were biphasic force-dependent,showing a'Catch-slip bond'transform in binding of A1 to A2 or A3.It suggested that under the low wall shear stresses,force could inhibit VWF activity through a catch bond mechanism,which enhanced the stability of the closed A1A2A3 conformation,but under high wall shear stresses,the force would enhance VWF activity through a slip bond mechanism,which promoted conformational transform of VWF-A from closed to extended one through reducing the stability of the closed A1A2A3 structure.Our results showed that the GOF mutant R1 308 L would down-regulate the binding affinity of A1 to A2,leading to a low barrier in opening of the closed VWF-A structure.In contrast,the LOF mutant G1324S would enhance the stability of the closed VWF-A conformation by up-regulating the binding affinity of A1 to A2,leading to inhibition of VWF activity.
文摘The characteristics of morphologies and changes of the microstructures in mechanically alloyed （MA） （m（Cr）∶m（C）∶m（Cr）=90∶1∶9） mixed powders were investigated by using SEM observation and X-ray diffraction （XRD） analyses. The results show that Cu, C and Cr mixed powders mechanically alloyed for 12 h forms the Cu-based super-saturated solid solution; after high energy ball-milling for 96 h the partial solute elements Cr and C precipitate, meanwhile, the mechanochemical reactions take place and the carbides Cr7C3 form. The composite powders after mechanically alloyed are fine and flat. The formed defects such as free surface, grain boundary, sub-grain boundary and dislocation decrease diffusion activity energy, which is the important reason for developing super-saturated solid solution and promoting the second phase precipitation.
基金supported by the National Natural Science Foundation of China ( 116272109, 11432006)
文摘Integrin activation,the transition from a low to a high affinity state,regulates the numerous cellular responses consequent to integrin engagement by extracellular matrix proteins.Kindlin proteins,play crucial roles in the integrin-signaling pathway by directly interacting with and activating integrins,which mediate the cell-extracellular matrix adhesion and signaling.As a widely distributed PTB domain protein and a major member of the kindlin family,kindlin2 interacts withβ3-tail,bridges talin-activated integrins to promote integrin aggregation,and enhances talin-induced integrin activation.Thus,kindlin2 is identified as a coactivator of integrins.Unlike talins,kindlin2 cannot directly alter the conformation of the integrin transmembrane helix and fail to activate integrin alone.Nevertheless,although it is widely accepted that kindlins and talins synergistically promote integrin activation,the underlying mechanism is unclear.Thus,the study of the force dissociation of the kindlin2/β3-tail complex and the conformation stabilization under different mechanical micro-environments should be of great significance for the further understanding of the structural basis of its synergistically activation of integrin.To reveal the molecular dynamics mechanism of interaction between kindlin2 andβ3-tail,we perform molecular dynamics(MD)simulations for this complex with different computing strategies interaction.In MD simulations,the available crystal structures of Kindlin-2/β3-tail complex(Protein Data Bank code 5XQ1)was downloaded from the PDB database.Two software packages,VMD for visualization and modeling and NAMD 2.13 for energy minimizations and MD simulations,were used here.The steadystate conformation of the complex was obtained from the equilibrium simulation.The dissociation event was observed by the constant velocity simulation,and the mechanical stability of the complex was observed by the constant force simulation.Our results showed that,during the equilibrium of the kindlin2-F3/β34ail complex,the residue MET612,LYS613 and TRP615 on the F3 domain of kindlin2 contributed to hydrogen-bonding with the corresponding residues onβ3 integrin.These bonds exhibit moderate or strong stability through steered molecular dynamics(SMD)simulation.During the constant velocity simulation,the complex exhibits a variety of unfolding pathways against tension applications,which are mainly distinguished by the disruption of hydrogen-bonds between the F3 domain a1/a2 helixes andβ1/β2 sheets.During the constant force simulation,the different phases of the composite force dissociation have different dissociation probabilities,which shows the biphasic force-dependent characteristics.And,the key residues in the pulling were recognized according not only to the number of interacting residue pairs,but also to their bond strength.Using molecular dynamics simulation,we showed the steady state of the kindlin2-F3/β3-tail complex under different tensile forces,and observe the dynamic process of molecular interaction.A possible underlying biophysical mechanism is that,the dissociation of Kindlin2-F3/β3-tail complex is biphasic force-dependent,and the conformations under different stretching states have different binding affinities.This study not only provides insights into the structural basis and mechanical regulation mechanisms of the kindlin/integrin interaction,in understanding in kindlin/integrin-related signaling in different cellular biological processes,but also provides new ideas for novel drug design and the treatment of related diseases.
基金supported by the National Natural Science Foundation of China ( 116272109, 11432006)
文摘CD40 receptor and its ligand CD40 L are the members of TNFR and TNF family,respectively.CD40 is expressed on antigen-presenting cells(APC)as a monomer,and would be dimerized upon its oligomerization through binding to its ligand CD40 L.Such dimer formation was shown to be essential for some CD40 signaling events,including phosphoinositide-3 kinase(PI-3 K)activation,the subsequent B7-2 upregulation and the production of IL-8.CD40 L is primarily expressed on activated T cells and activated platelets,and acts as an important costimulatory molecule especially on T follicular helper cells.It is believed that CD40-CD40 L interaction deeply involved in many immune events for the host defense against pathogens and cancer.CD40-CD40 L interaction mediates intracellular signaling along different pathways,such as the canonical and noncanonical nuclear factorκB pathway,mitogen-activated protein kinases,phosphatidylinositol-3 kinase(PI3K)and the phospholipase Cy pathway,is necessary for successful adaptive immune responses mainly relevant to development of CD8+CTLs,and is deeply involved in cross-talks among T cell,B cell,tumor cell,platelet and etc.However,interaction of CD40 with platelet CD40 L would occur in hemodynamics environments,and the regulation of force on this interaction remain unclear.Besides polymerization of CD40,the hemodynamic environment could be believed to regulate interaction between CD40 and its ligand.To reveal the mechanical regulation mechanism of interaction between CD40 L and CD40,atomic force microscope(AFM)was used in the adhesion frequency assay for measuring two-dimensional(2D)kinetics of CD40 L with CD40(the monomer and its dimer)at zero-force and force-clamp assay for measuring single-bond lifetimes of CD40 L/CD40 complex in a range of forces at single-molecule level.AFM cantilever probes were functionalized by incubation with CD40 L in buffer,and soaked in phosphate-buffered saline(PBS)containing 1%bovine serum albumin(BSA)to block nonspecific binding.Polystyrene dishes were coated with CD40(the monomer and its dimer)and then filled with PBS containing 1%BSA.Our data showed that the adhesion frequency of CD40 L with CD40 monomer and its dimer was 0.10 and0. 15,respectively,showing a higher affinity of CD40 L to CD40 dimer rather than its monomer;the rupture force for CD40 L dissociating from CD40 monomer and its dimer was 23 and 31pN,respectively,demonstrating a higher mechanical strength of complex of CD40 L with CD40 dimer instead of monomer;and increasing tension will prolong first and then shorten the bond lifetime of CD40 L-CD40 complex with force threshold of about 1OpN,and dimerization of CD40 had significantly increased enhanced the bond lifetime of CD40 L-CD40 complex,showing a dimerization-enhanced affinity of CD40 to CD40 L and'Catch-Slip'bond mechanism of CD40 L-CD40 interaction.These results suggest that,through upregulating the binding affinity with CD40 L,the polymerization of CD40 stabilizes the linkers and thereby communication between cell and cell,so does the force being small than the force threshold Both CD40 polymerization-and'Catch bond'-induced enhancement of affinity of CD40 with CD40 L may be necessary for stable cross talks between two cells.This finding may be useful for understanding CD40/CD40 L-induced events importantly in physiological or pathological processes at molecular and cellular level.
基金supported by the National Natural Science Foundation of China ( 116272109, 11432006)
文摘As a key regulator of immune response,CD40 L is usually associated with chronic disease-related inflammation,autoimmune diseases and malignant diseases.Receptor recognition of platelet CD40 L is the initial event that mediates platelet aggregation and leukocyte immune response.Unlike soluble CD40 L,the interaction between transmembrane platelet CD40 L and its receptors occurs within the cell junction surface,usually,in a physiological and pathological high blood flow shear stress environment.This two-dimensional reaction kinetics should be a mechano-chemical coupling process.In addition to its classical receptor CD40,CD40 L also binds to receptorα5β1,CD40 L can bind to the resting state of integrinα5β1,but the mechanical regulation mechanism of integrinα5β1 activation under fluid shear stress remains unclear.We assume that the force can promote CD40 L-inducedα5β1 activation.To check this hypothesis,we performed flow chamber experiment to investigate interaction of CD40 L andα5β1.In experiments,the bottom of the flow chamber is functionalized by a suitable concentration of CD40 L,and the fiber spheres of 6μm diameter was coated withα5β1.The selection of CD40 L concentration was based on the observation that as many tether events ofα5β1-coated spheres as possible were observed rather than stable adhesion events of these spheres.Theα5β1-coated sphere suspension was poured over the CD40 L-coated substrates in the flow chamber under different shear rates.A high-speed camera was used to observe and record tether events of fiber spheres at a rate of 100 frames per second.According to our affinity state transition model for integrin,the data were analyzed to obtain the rate of integrin activation and its mechanical regulation characteristics.Our results demonstrated that the interaction betweenα5β1 and CD40 L is biphasic force-dependent,showing mechano-chemical regulation mechanism of'Catch-slip bond'transition.The affinity jumping model was well fitted with the data obtained from flow chamber experiment at various wall shear stresses.We found that,CD40 L ligation-induced jumping ofα5β1 affinity state from low to medium(or high)one will occur within 0.5-1.0 second,resulting in prolonging of bond lifetimes.And,frequency distribution of the tether events number with tether lifetime under each force,exhibits obvious doublet peaks,one within 0.5-1 s and second within 1.5-2.5 s,indicating theα5β1 affinity state transform from low to high one.The probability distribution of the tether lifetime under different shear forces are not linear,and exists a turning point,which shows that the rate ofα5β1 dissociation from CD40 L is fast first,and then become slow,showing a force-induced conformation transformation of the integrinα5β1 from low affinity state to high affinity one.Our findings suggest that,the continuous force stimulation will quickly cause the affinity state change of integrinα5β1. The dissociation rate of theα5β1/CD40 L complex decreases first and then increases with wall shear stress,exhibiting a'Catch-slip bond'transformation of interaction betweenα5β1-CD40 L.This mechanical regulation mechanism exists in interaction of CD40 L not only toα5β1 at low affinity state but also to one at high affinity state.Our results should be useful in understanding the mechanical regulation mechanism of a5β1-CD40 L interaction-mediated cellular immune response and inflammatory processes.
