Homodimeric prodrug-based self-assembled nanoparticles,with carrier-free structure and ultrahigh drug loading,is drawing more and more attentions.Homodimeric prodrugs are composed of two drug molecules and a pivotal l...Homodimeric prodrug-based self-assembled nanoparticles,with carrier-free structure and ultrahigh drug loading,is drawing more and more attentions.Homodimeric prodrugs are composed of two drug molecules and a pivotal linkage.The influence of the linkages on the self-assembly,in vivo fate and antitumor activity of homodimeric prodrugs is the focus of research.Herein,three docetaxel(DTX)homodimeric prodrugs are developed using different lengths of diselenide bond-containing linkages.Interestingly,compared with the other two linkages,the longest diselenide bond-containing linkage could facilitate the self-delivery of DTX prodrugs,thus improving the stability,circulation time and tumor targeting of prodrug nanoassemblies.Besides,the extension of linkages reduces the redox-triggered drug release and cytotoxicity of prodrug nanoassemblies in tumor cells.Although the longest diselenide bond-containing prodrug nanoassemblies possessed the lowest cytotoxicity to 4T1 cells,their stable nanostructure maintained intact during circulation and achieve the maximum accumulation of DTX in tumor cells,which finally“turned the table”.Our study illustrates the crucial role of linkages in homodimeric prodrugs,and gives valuable proposal for the development of advanced nano-DDS for cancer treatment.展开更多
Amongst endonuclease, the homodimeric variety is found in many prokaryotes for processing of the introns out from pre-RNAs. But as the variety and the complexity of introns rise with evolution, do the homodimeric endo...Amongst endonuclease, the homodimeric variety is found in many prokaryotes for processing of the introns out from pre-RNAs. But as the variety and the complexity of introns rise with evolution, do the homodimeric endonuclease adapt to the changes? The correlations between evolving pre-RNAs and adapting homodimeric endonuclease in lower prokaryotes is investigated in this paper. First, we construct and observe the appearance of a long branch in the phylogeny based on homodimeric endonuclease. To appreciate the finer aspects of accelerating evolution near this long branch, we delve deeper into the pre-RNA substrates of the endonuclease. Computational evidence of an as-yet-unreported noncoding RNA gene then emerges from this study. The capabilities of homodimeric endonuclease and the complexities of its pre-RNA substrates appear to evolve in steps together.展开更多
Isoflavones which mainly distributed in leguminous plants have plenty of health benefits.Isoflavone synthase(IFS)is a membrane-associated cytochrome P450 enzyme(CYP450)which carries out the unique aryl-ring migration ...Isoflavones which mainly distributed in leguminous plants have plenty of health benefits.Isoflavone synthase(IFS)is a membrane-associated cytochrome P450 enzyme(CYP450)which carries out the unique aryl-ring migration and hydroxylation.So far,few crystal structures of plant P450s have been obtained.We determined the crystal structure of IFS from Medicago truncatula at 1.9 by MAD method using a selenomethionine substituted crystal and conducted molecular docking and mutagenesis study.The structure of IFS complexed with imidazole exhibits the helix Iα-loop-helix Iβmotif which corresponds to helix I of other P 450s.Compared with structures of common P450s,IFS/imidazole structure contains an extra domain,i.e.,theγ-domain.The structure reveals a homodimer in which theγ-domain of one molecule interacts with theβ-domain of another.The plane of heme group makes an angle of approximately 40°with the helix Iα-loop-helix Iβmotif.Molecular docking combined with mutagenesis study suggested that Trp-128 and Asp-300 might play important roles in substrate binding and recognition.Phe-301,Ser-303 and Gly-305 from the helix Iα-loop-helix Iβmotif may play important roles in the aryl-ring migration.These novel structural features reveal insights into the unique reaction mechanism of IFS and provide a basis for engineering IFS in leguminous crops for health purpose.展开更多
The evolution of severe acute respiratory syndrome coronavirus 2(SARS-CoV-2)has resulted in mutations not only in the spike protein,aiding immune evasion,but also in the NSP3/4/6 proteins,crucial for regulating double...The evolution of severe acute respiratory syndrome coronavirus 2(SARS-CoV-2)has resulted in mutations not only in the spike protein,aiding immune evasion,but also in the NSP3/4/6 proteins,crucial for regulating double-membrane vesicle(DMV)formation.However,the functional consequences of these NSP3/4/6 mutations remain poorly understood.In this study,a systematic analysis was conducted to investigate the evolutionary patterns of NSP3/4/6 mutations and their impact on DMV formation.The findings revealed that the NSP4 T492I mutation,a prevalent mutation found in all Delta and Omicron sub-lineages,notably enhances DMV formation.Mechanistically,the NSP4 T492I mutation enhances its homodimerization,leading to an increase in the size of puncta induced by NSP3/4,and also augments endoplasmic reticulum(ER)membrane curvature,resulting in a higher DMV density per fluorescent puncta.This study underscores the significance of the NSP4 T492I mutation in modulating DMV formation,with potential implications for the transmission dynamics of SARS-CoV-2.It contributes valuable insights into how these mutations impact viral replication and pathogenesis.展开更多
Cytokinins(CKs),primarily trans-zeatin(tZ)and isopentenyladenine(iP)types,play critical roles in plant growth,development,and various stress responses.Long-distance transport of tZ-type CKs meidated by Arabidopsis ATP...Cytokinins(CKs),primarily trans-zeatin(tZ)and isopentenyladenine(iP)types,play critical roles in plant growth,development,and various stress responses.Long-distance transport of tZ-type CKs meidated by Arabidopsis ATP-binding cassette transporter subfamily G14(AtABCG14)has been well studied;however,less is known about the biochemical properties of AtABCG14 and its transporter activity toward iP-type CKs.Here we reveal the biochemical properties of AtABCG14 and provide evidence that it is also required for long-distance transport of iP-type CKs.AtABCG14 formed homodimers in human(Homo sapiens)HEK293T,tobacco(Nicotiana tabacum),and Arabidopsis cells.Transporter activity assays of AtABCG14 in Arabidopsis,tobacco,and yeast(Saccharomyces cerevisiae)showed that AtABCG14 may directly transport multiple CKs,including iP-and tZ-type species.AtABCG14 expression was induced by iP in a tZ-type CKdeficient double mutant(cypDM)of CYP735A1 and CYP735A2.The atabcg14 cypDM triple mutant exhibited stronger CK-deficiency phenotypes than cypDM.Hormone profiling,reciprocal grafting,and 2H6-iP isotope tracer experiments showed that root-to-shoot and shoot-to-root long-distance transport of iP-type CKs were suppressed in atabcg14 cypDM and atabcg14.These results suggest that AtABCG14 participates in three steps of the circular long-distance transport of iP-type CKs:xylem loading in the root for shootward transport,phloem unloading in the shoot for shoot distribution,and phloemunloading in the root for root distribution.We found that AtABCG14 displays transporter activity toward multiple CK species and revealed its versatile roles in circular long-distance transport of iP-type CKs.These findings provide newinsights into the transport mechanisms of CKs and other plant hormones.展开更多
Bispecific antibodies hold significant potential as next-generation biotherapeutics owing to their ability to simultaneously bind to two targets.However,the development of bispecific antibodies as biotherapeutics has ...Bispecific antibodies hold significant potential as next-generation biotherapeutics owing to their ability to simultaneously bind to two targets.However,the development of bispecific antibodies as biotherapeutics has been hindered by the high levels of byproducts produced,including both high molecular weight and low molecular weight variants.In addition,the inevitable expression of homodimers in host cells presents further obstacles to the commercial development of bispecific antibodies as therapeutics.These byproducts,which share similar physicochemical properties with the target,pose several challenges for downstream purification processes.In this study,we present a non-protein A purification platform that employ a one-step polishing chromatography to purify bispecific antibodies.Mixed-mode Capto adhere resin was used to capture the target protein at pH 7.90±0.10,followed by anion exchange chromatography as a polishing step.Overall,the results of this two-step chromatography purification method demonstrated at final product purity of 98%as assessed by size-exclusion high-performance liquid chromatography(SEC-HPLC)and 98%by reversed-phase-high-performance liquid chromatography(RP-HPLC),with residual host cell proteins controlled at 10 ppm and an excellent recovery rate of approximately 60%.This study presents a non-protein A capture platform,offering a simplified,streamlined,and competitive alternative to conventional affinity chromatography.展开更多
[Objective] The aim was to investigate the dimer formation between the movement proteins（MP）in barely yellow dwarf virus by using the technology of bimolecular fluorescence complementation technology and to further ...[Objective] The aim was to investigate the dimer formation between the movement proteins（MP）in barely yellow dwarf virus by using the technology of bimolecular fluorescence complementation technology and to further study the relationship between MP homodimerization and viral movement.[Method] The DNA sequence of bimolecular fluorescent complementary vector containing cloning multiple cloning sites,35S promoter and terminator was cloned into the expression vector pCAMBIA1300,which replicates at a higher copy number in E.coli.Then,the BYDV-MP gene fragment was amplified in the presence of the whole BYDV-PAV cDNA sequence as template and the primers designed according to the BYDV-MP gene sequence from GenBank,cloned into the modified bimolecular fluorescent complementary vectors pCAMBIA1300-NE and pCAMBIA1300-CE.The resulting vectors were transformed into Agrobacterium by electroporation method and infiltrated into the tobacco leaf.Protein interactions were observed under fluorescence microscope.[Result] Yellow fluorescence could be viewed in the leaves co-infiltrated with Agrobacterium carrying pCAMBIA1300NE-MP and pCAMBIA1300CE-MP at 2-5 d post-infiltration,while yellow fluorescence could not be observed in negative control groups.[Conclusion] BYDV-MP formed homodimers in plant cells.The results can provide theoretical basis for further in-depth research about the movement process and mechanism of BYDV.展开更多
Infrared multiple photon dissociation （IRMPD） spectroscopic and theoretical studies of protonated homodimers of amino acids generated by electrospray ionization in the gas phase have been reviewed. Results show that...Infrared multiple photon dissociation （IRMPD） spectroscopic and theoretical studies of protonated homodimers of amino acids generated by electrospray ionization in the gas phase have been reviewed. Results show that proton affinity （PA） may be applied as a probe to predict their structural type： salt- bridged or charge-solvated. Proline can be viewed as a reference. For an amino acid with a PA value higher than that of proline, the most stable conformation of its protonated homodimer tends to prefer salt- bridged conformation; otherwise, charge-solvated conformation is expected to be the most stable. However, side chain effects may cause the inaccuracy in structural determination due to the strong interactions with the charge, which makes the charge-solvated structure more stable even for species with high PA values. Temperature effect on distribution of different isomers is also very important. In lots of cases, the coexistence of multiple isomers is general, which makes the explanation of an overall IRMPD spectrum difficult. So a statistical view on the distribution of optimized isomers is very helpful.展开更多
N-Methyl-D-aspartate receptors（NMDARs） play a central role in various physiological and pathological processes in the central nervous system.And they are commonly composed of four subunits,two GluN1 subunits and two...N-Methyl-D-aspartate receptors（NMDARs） play a central role in various physiological and pathological processes in the central nervous system.And they are commonly composed of four subunits,two GluN1 subunits and two GluN2 or GluN3 subunits.The different subunit compositions make NMDARs a heterogeneous population with distinct electrophysiological and pharmacological properties and thus with different abilities to conduct neuronal activities.The subunit composition,assembly process,and final structure of assembled NMDARs have been studied for years but no consensus has been achieved.In this study,we investigated the role of the amino terminal domain（ATD） of GluN2A in regulating NMDAR assembly.The ATD of GluN2A was first expressed in heterogeneous cells and the homodimer formation was investigated by fluorescent resonance energy transfer and non-reducing SDS- PAGE electrophoresis.Each of the three cysteine residues located in the ATD was mutated into alanine,and the homodimerization of the ATD or GluN2A,as well as the heteromeric assembly of NMDARs was assessed by non-reducing SDS- PAGE electrophoresis,co-immunoprecipitation and immunocytochemistry.We found that two cysteine residues,C87 and C320,in the ATD of the GluN2A subunit were required for the formation of disulfide bonds and GluN2A ATD homodimers.Furthermore,the disruption of GluN2A ATD domain dimerization had no influence on the assembly and surface expression of NMDARs.These results suggest that the two ATD domains of GluN2A are structurally adjacent in fully-assembled NMDARs.However,unlike GluN1,the homomerization of the ATD domain of GluN2A is not required for the assembly of NMDARs,implying that GluN2A and GluN1 play unequal roles in NMDAR assembly.展开更多
Chemotherapy has occupied the critical position in cancer therapy,especially towards the post-operative,advanced,recurrent,and metastatic tumors.Paclitaxel(PTX)-based formulations have been widely used in clinical pra...Chemotherapy has occupied the critical position in cancer therapy,especially towards the post-operative,advanced,recurrent,and metastatic tumors.Paclitaxel(PTX)-based formulations have been widely used in clinical practice,while the therapeutic effect is far from satisfied due to off-target toxicity and drug resistance.The caseless multi-components make the preparation technology complicated and aggravate the concerns with the excipients-associated toxicity.The self-assembled PTX nanoparticles possess a high drug content and could incorporate various functional molecules for enhancing the therapeutic index.In this work,we summarize the self-assembly strategy for diverse nanodrugs of PTX.Then,the advancement of nanodrugs for tumor therapy,especially emphasis on monochemotherapy,combinational therapy,and theranostics,have been outlined.Finally,the challenges and potential improvements have been briefly spotlighted.展开更多
MRG proteins are conserved during evolution in fungi, flies, mammals and plants, and they can exhibit diversified functions. The animal MRGs were found to form various complexes to activate gene expression. Plant MRGI...MRG proteins are conserved during evolution in fungi, flies, mammals and plants, and they can exhibit diversified functions. The animal MRGs were found to form various complexes to activate gene expression. Plant MRGI/2 and MRG702 were reported to be involved in the regulation of flowering time via binding to H3K36me3- marked flowering genes. Herein, we determined the crystal structure of MRG701 chromodomain （MRG701CD）. MRG701co forms a novel dimerization fold both in crystal and in solution. Moreover, we found that the dimerization of MRG chromodomains is conserved in green plants. Our findings may provide new insights into the mechanism of MRGs in regulation of gene expression in green plants.展开更多
Nascent polypeptide associated complex(NAC)and its two isolated subunits,αNAC and βNAC,play important roles in nascent peptide targeting.We determined a 1.9Åresolution crystal structure of the interaction core ...Nascent polypeptide associated complex(NAC)and its two isolated subunits,αNAC and βNAC,play important roles in nascent peptide targeting.We determined a 1.9Åresolution crystal structure of the interaction core of NAC heterodimer and a 2.4Åresolution crystal structure ofαNAC NAC domain homodimer.These structures provide detailed information of NAC heterodimerization and αNAC homodimerization.We found that the NAC domains of αNAC and βNAC share very similar folding despite of their relative low identity of amino acid sequences.Furthermore,different electric charge distributions of the two subunits at the NAC interface provide an explanation to the observation that the heterodimer of NAC complex is more stable than the single subunit homodimer.In addition,we successfully built a βNAC NAC domain homodimer model based on homologous modeling,suggesting that NAC domain dimerization is a general property of the NAC family.These 3D structures allow further studies on structurefunction relationship of NAC.展开更多
Sometimes crystals cannot diffract X-rays beyond 3.0Åresolution due to the intrinsic flexibility associated with the protein.Low resolution diffraction data not only pose a challenge to structure determination,bu...Sometimes crystals cannot diffract X-rays beyond 3.0Åresolution due to the intrinsic flexibility associated with the protein.Low resolution diffraction data not only pose a challenge to structure determination,but also hamper interpretation of mechanistic details.Crystals of a 25.6 kDa non-Pfam,hypothetical protein,PF2046,diffracted X-rays to 3.38Åresolution.A combination of SeMet derived heavy atom positions with multiple cycles of B-factor sharpening,multi-crystal averaging,restrained refinement followed by manual inspection of electron density and model building resulted in a final model with a R value of 23.5(R_(free)=24.7).The asymmetric unit was large and consisted of six molecules arranged as a homodimer of trimers.Analysis of the structure revealed the presence of a RNA binding domain suggesting a role for PF2046 in the processing of nucleic acids.展开更多
基金This work was supported by China Postdoctoral Innovative Talents Support Program(no.BX20190219)China Postdoctoral Science Foundation(no.2019M661134)National Natural Science Foundation of China(no.81872816).
文摘Homodimeric prodrug-based self-assembled nanoparticles,with carrier-free structure and ultrahigh drug loading,is drawing more and more attentions.Homodimeric prodrugs are composed of two drug molecules and a pivotal linkage.The influence of the linkages on the self-assembly,in vivo fate and antitumor activity of homodimeric prodrugs is the focus of research.Herein,three docetaxel(DTX)homodimeric prodrugs are developed using different lengths of diselenide bond-containing linkages.Interestingly,compared with the other two linkages,the longest diselenide bond-containing linkage could facilitate the self-delivery of DTX prodrugs,thus improving the stability,circulation time and tumor targeting of prodrug nanoassemblies.Besides,the extension of linkages reduces the redox-triggered drug release and cytotoxicity of prodrug nanoassemblies in tumor cells.Although the longest diselenide bond-containing prodrug nanoassemblies possessed the lowest cytotoxicity to 4T1 cells,their stable nanostructure maintained intact during circulation and achieve the maximum accumulation of DTX in tumor cells,which finally“turned the table”.Our study illustrates the crucial role of linkages in homodimeric prodrugs,and gives valuable proposal for the development of advanced nano-DDS for cancer treatment.
文摘Amongst endonuclease, the homodimeric variety is found in many prokaryotes for processing of the introns out from pre-RNAs. But as the variety and the complexity of introns rise with evolution, do the homodimeric endonuclease adapt to the changes? The correlations between evolving pre-RNAs and adapting homodimeric endonuclease in lower prokaryotes is investigated in this paper. First, we construct and observe the appearance of a long branch in the phylogeny based on homodimeric endonuclease. To appreciate the finer aspects of accelerating evolution near this long branch, we delve deeper into the pre-RNA substrates of the endonuclease. Computational evidence of an as-yet-unreported noncoding RNA gene then emerges from this study. The capabilities of homodimeric endonuclease and the complexities of its pre-RNA substrates appear to evolve in steps together.
文摘Isoflavones which mainly distributed in leguminous plants have plenty of health benefits.Isoflavone synthase(IFS)is a membrane-associated cytochrome P450 enzyme(CYP450)which carries out the unique aryl-ring migration and hydroxylation.So far,few crystal structures of plant P450s have been obtained.We determined the crystal structure of IFS from Medicago truncatula at 1.9 by MAD method using a selenomethionine substituted crystal and conducted molecular docking and mutagenesis study.The structure of IFS complexed with imidazole exhibits the helix Iα-loop-helix Iβmotif which corresponds to helix I of other P 450s.Compared with structures of common P450s,IFS/imidazole structure contains an extra domain,i.e.,theγ-domain.The structure reveals a homodimer in which theγ-domain of one molecule interacts with theβ-domain of another.The plane of heme group makes an angle of approximately 40°with the helix Iα-loop-helix Iβmotif.Molecular docking combined with mutagenesis study suggested that Trp-128 and Asp-300 might play important roles in substrate binding and recognition.Phe-301,Ser-303 and Gly-305 from the helix Iα-loop-helix Iβmotif may play important roles in the aryl-ring migration.These novel structural features reveal insights into the unique reaction mechanism of IFS and provide a basis for engineering IFS in leguminous crops for health purpose.
基金supported by the National Natural Science Foundation of China(92469107 to Z.Li)the R&D Program of Guangzhou National Laboratory(ZL-SRPG2200205 to Z.Li)the Guangdong Province High-level Talent Youth Project(2021QN02Y939 to Z.Li).
文摘The evolution of severe acute respiratory syndrome coronavirus 2(SARS-CoV-2)has resulted in mutations not only in the spike protein,aiding immune evasion,but also in the NSP3/4/6 proteins,crucial for regulating double-membrane vesicle(DMV)formation.However,the functional consequences of these NSP3/4/6 mutations remain poorly understood.In this study,a systematic analysis was conducted to investigate the evolutionary patterns of NSP3/4/6 mutations and their impact on DMV formation.The findings revealed that the NSP4 T492I mutation,a prevalent mutation found in all Delta and Omicron sub-lineages,notably enhances DMV formation.Mechanistically,the NSP4 T492I mutation enhances its homodimerization,leading to an increase in the size of puncta induced by NSP3/4,and also augments endoplasmic reticulum(ER)membrane curvature,resulting in a higher DMV density per fluorescent puncta.This study underscores the significance of the NSP4 T492I mutation in modulating DMV formation,with potential implications for the transmission dynamics of SARS-CoV-2.It contributes valuable insights into how these mutations impact viral replication and pathogenesis.
基金the National Natural Science Foundation of China(31470370 to K.Z.and 32100270 to J.Z.)the Natural Science Foundation of Zhejiang Province(LY22C020003 to J.Z.)C.-J.L.was supported by the DOE Office of Basic Energy Sciences,specifically through the Physical Biosciences Program of the Chemical Sciences,Geosciences and Biosciences Division,under contract DESC0012704.
文摘Cytokinins(CKs),primarily trans-zeatin(tZ)and isopentenyladenine(iP)types,play critical roles in plant growth,development,and various stress responses.Long-distance transport of tZ-type CKs meidated by Arabidopsis ATP-binding cassette transporter subfamily G14(AtABCG14)has been well studied;however,less is known about the biochemical properties of AtABCG14 and its transporter activity toward iP-type CKs.Here we reveal the biochemical properties of AtABCG14 and provide evidence that it is also required for long-distance transport of iP-type CKs.AtABCG14 formed homodimers in human(Homo sapiens)HEK293T,tobacco(Nicotiana tabacum),and Arabidopsis cells.Transporter activity assays of AtABCG14 in Arabidopsis,tobacco,and yeast(Saccharomyces cerevisiae)showed that AtABCG14 may directly transport multiple CKs,including iP-and tZ-type species.AtABCG14 expression was induced by iP in a tZ-type CKdeficient double mutant(cypDM)of CYP735A1 and CYP735A2.The atabcg14 cypDM triple mutant exhibited stronger CK-deficiency phenotypes than cypDM.Hormone profiling,reciprocal grafting,and 2H6-iP isotope tracer experiments showed that root-to-shoot and shoot-to-root long-distance transport of iP-type CKs were suppressed in atabcg14 cypDM and atabcg14.These results suggest that AtABCG14 participates in three steps of the circular long-distance transport of iP-type CKs:xylem loading in the root for shootward transport,phloem unloading in the shoot for shoot distribution,and phloemunloading in the root for root distribution.We found that AtABCG14 displays transporter activity toward multiple CK species and revealed its versatile roles in circular long-distance transport of iP-type CKs.These findings provide newinsights into the transport mechanisms of CKs and other plant hormones.
文摘Bispecific antibodies hold significant potential as next-generation biotherapeutics owing to their ability to simultaneously bind to two targets.However,the development of bispecific antibodies as biotherapeutics has been hindered by the high levels of byproducts produced,including both high molecular weight and low molecular weight variants.In addition,the inevitable expression of homodimers in host cells presents further obstacles to the commercial development of bispecific antibodies as therapeutics.These byproducts,which share similar physicochemical properties with the target,pose several challenges for downstream purification processes.In this study,we present a non-protein A purification platform that employ a one-step polishing chromatography to purify bispecific antibodies.Mixed-mode Capto adhere resin was used to capture the target protein at pH 7.90±0.10,followed by anion exchange chromatography as a polishing step.Overall,the results of this two-step chromatography purification method demonstrated at final product purity of 98%as assessed by size-exclusion high-performance liquid chromatography(SEC-HPLC)and 98%by reversed-phase-high-performance liquid chromatography(RP-HPLC),with residual host cell proteins controlled at 10 ppm and an excellent recovery rate of approximately 60%.This study presents a non-protein A capture platform,offering a simplified,streamlined,and competitive alternative to conventional affinity chromatography.
基金Supported by National Natural Science Foundation of China(30870109)~~
文摘[Objective] The aim was to investigate the dimer formation between the movement proteins（MP）in barely yellow dwarf virus by using the technology of bimolecular fluorescence complementation technology and to further study the relationship between MP homodimerization and viral movement.[Method] The DNA sequence of bimolecular fluorescent complementary vector containing cloning multiple cloning sites,35S promoter and terminator was cloned into the expression vector pCAMBIA1300,which replicates at a higher copy number in E.coli.Then,the BYDV-MP gene fragment was amplified in the presence of the whole BYDV-PAV cDNA sequence as template and the primers designed according to the BYDV-MP gene sequence from GenBank,cloned into the modified bimolecular fluorescent complementary vectors pCAMBIA1300-NE and pCAMBIA1300-CE.The resulting vectors were transformed into Agrobacterium by electroporation method and infiltrated into the tobacco leaf.Protein interactions were observed under fluorescence microscope.[Result] Yellow fluorescence could be viewed in the leaves co-infiltrated with Agrobacterium carrying pCAMBIA1300NE-MP and pCAMBIA1300CE-MP at 2-5 d post-infiltration,while yellow fluorescence could not be observed in negative control groups.[Conclusion] BYDV-MP formed homodimers in plant cells.The results can provide theoretical basis for further in-depth research about the movement process and mechanism of BYDV.
基金supported by the National Natural Science Foundation of China(Nos.21475065,21627801 and 11704280)
文摘Infrared multiple photon dissociation （IRMPD） spectroscopic and theoretical studies of protonated homodimers of amino acids generated by electrospray ionization in the gas phase have been reviewed. Results show that proton affinity （PA） may be applied as a probe to predict their structural type： salt- bridged or charge-solvated. Proline can be viewed as a reference. For an amino acid with a PA value higher than that of proline, the most stable conformation of its protonated homodimer tends to prefer salt- bridged conformation; otherwise, charge-solvated conformation is expected to be the most stable. However, side chain effects may cause the inaccuracy in structural determination due to the strong interactions with the charge, which makes the charge-solvated structure more stable even for species with high PA values. Temperature effect on distribution of different isomers is also very important. In lots of cases, the coexistence of multiple isomers is general, which makes the explanation of an overall IRMPD spectrum difficult. So a statistical view on the distribution of optimized isomers is very helpful.
基金supported by grants from the National Basic Research Development Program of China(2010CB912002)the National Natural Science Foundation of China(30730038 and 81171164)
文摘N-Methyl-D-aspartate receptors（NMDARs） play a central role in various physiological and pathological processes in the central nervous system.And they are commonly composed of four subunits,two GluN1 subunits and two GluN2 or GluN3 subunits.The different subunit compositions make NMDARs a heterogeneous population with distinct electrophysiological and pharmacological properties and thus with different abilities to conduct neuronal activities.The subunit composition,assembly process,and final structure of assembled NMDARs have been studied for years but no consensus has been achieved.In this study,we investigated the role of the amino terminal domain（ATD） of GluN2A in regulating NMDAR assembly.The ATD of GluN2A was first expressed in heterogeneous cells and the homodimer formation was investigated by fluorescent resonance energy transfer and non-reducing SDS- PAGE electrophoresis.Each of the three cysteine residues located in the ATD was mutated into alanine,and the homodimerization of the ATD or GluN2A,as well as the heteromeric assembly of NMDARs was assessed by non-reducing SDS- PAGE electrophoresis,co-immunoprecipitation and immunocytochemistry.We found that two cysteine residues,C87 and C320,in the ATD of the GluN2A subunit were required for the formation of disulfide bonds and GluN2A ATD homodimers.Furthermore,the disruption of GluN2A ATD domain dimerization had no influence on the assembly and surface expression of NMDARs.These results suggest that the two ATD domains of GluN2A are structurally adjacent in fully-assembled NMDARs.However,unlike GluN1,the homomerization of the ATD domain of GluN2A is not required for the assembly of NMDARs,implying that GluN2A and GluN1 play unequal roles in NMDAR assembly.
基金the financial support from the National Natural Science Foundation of China(Nos.52203199 and 51973214)the Natural Science Foundation of Jilin Province(No.YDZJ202101ZYTS013)。
文摘Chemotherapy has occupied the critical position in cancer therapy,especially towards the post-operative,advanced,recurrent,and metastatic tumors.Paclitaxel(PTX)-based formulations have been widely used in clinical practice,while the therapeutic effect is far from satisfied due to off-target toxicity and drug resistance.The caseless multi-components make the preparation technology complicated and aggravate the concerns with the excipients-associated toxicity.The self-assembled PTX nanoparticles possess a high drug content and could incorporate various functional molecules for enhancing the therapeutic index.In this work,we summarize the self-assembly strategy for diverse nanodrugs of PTX.Then,the advancement of nanodrugs for tumor therapy,especially emphasis on monochemotherapy,combinational therapy,and theranostics,have been outlined.Finally,the challenges and potential improvements have been briefly spotlighted.
文摘MRG proteins are conserved during evolution in fungi, flies, mammals and plants, and they can exhibit diversified functions. The animal MRGs were found to form various complexes to activate gene expression. Plant MRGI/2 and MRG702 were reported to be involved in the regulation of flowering time via binding to H3K36me3- marked flowering genes. Herein, we determined the crystal structure of MRG701 chromodomain （MRG701CD）. MRG701co forms a novel dimerization fold both in crystal and in solution. Moreover, we found that the dimerization of MRG chromodomains is conserved in green plants. Our findings may provide new insights into the mechanism of MRGs in regulation of gene expression in green plants.
基金This work was supported by the National Natural Science Foundation of China(grant No.30730022)the National Basic Research Program(973 Program)(grant Nos.2006CB806503 and 2007CB914304)+1 种基金the National Programs for High Technology Research and Development Program(863 Program)(grant Nos.2006AA02A322 and 2006AA020502)the CAS(China)grant KSCX2-YW-R-05 to Z.R.
文摘Nascent polypeptide associated complex(NAC)and its two isolated subunits,αNAC and βNAC,play important roles in nascent peptide targeting.We determined a 1.9Åresolution crystal structure of the interaction core of NAC heterodimer and a 2.4Åresolution crystal structure ofαNAC NAC domain homodimer.These structures provide detailed information of NAC heterodimerization and αNAC homodimerization.We found that the NAC domains of αNAC and βNAC share very similar folding despite of their relative low identity of amino acid sequences.Furthermore,different electric charge distributions of the two subunits at the NAC interface provide an explanation to the observation that the heterodimer of NAC complex is more stable than the single subunit homodimer.In addition,we successfully built a βNAC NAC domain homodimer model based on homologous modeling,suggesting that NAC domain dimerization is a general property of the NAC family.These 3D structures allow further studies on structurefunction relationship of NAC.
基金This work was funded by the Ministry of Science and Technology of China(Grant Nos.2006AA02A316,2009DFB30310 and 2006CB910901)the National Natural Science Foundation of China(Grants Nos.30670427 and 30721003)+1 种基金the Ministry of Health of China(Grant No.2008ZX10404)CAS Research Grant(No.KSCX2-YW-R-127 and INFO-115-D01-2009).
文摘Sometimes crystals cannot diffract X-rays beyond 3.0Åresolution due to the intrinsic flexibility associated with the protein.Low resolution diffraction data not only pose a challenge to structure determination,but also hamper interpretation of mechanistic details.Crystals of a 25.6 kDa non-Pfam,hypothetical protein,PF2046,diffracted X-rays to 3.38Åresolution.A combination of SeMet derived heavy atom positions with multiple cycles of B-factor sharpening,multi-crystal averaging,restrained refinement followed by manual inspection of electron density and model building resulted in a final model with a R value of 23.5(R_(free)=24.7).The asymmetric unit was large and consisted of six molecules arranged as a homodimer of trimers.Analysis of the structure revealed the presence of a RNA binding domain suggesting a role for PF2046 in the processing of nucleic acids.
