Mammalian mitochondrial electron transport chain complexes are the most important and complicated protein machinery in mitochondria.Although this system has been studied for more than a century,its composition and mol...Mammalian mitochondrial electron transport chain complexes are the most important and complicated protein machinery in mitochondria.Although this system has been studied for more than a century,its composition and molecular mechanism are still largely unknown.Here we report the high-resolution cryo-electron microscopy(Cryo-EM)structures of porcine respiratory chain megacomplex-Ⅰ_(2)Ⅲ_(2)Ⅳ_(2)(MCⅠ_(2)Ⅲ_(2)Ⅳ_(2))in five different conformations,including State 1,State 2,Mid 1,Mid 2,and Mid 3.High-resolution Cryo-EM imaging,combined with super-resolution gated stimulated emission depletion microscopy(gSTED),strongly supports the formation of MCⅠ_(2)Ⅲ_(2)Ⅳ_(2)in live cells.Each MCⅠ_(2)Ⅲ_(2)Ⅳ_(2)structure contains 141 subunits(70 different kinds of peptides,2.9 MDa)in total with 240 transmembrane helices.The mutual influence among CⅠ,CⅢ,and CⅣshown in the MCⅠ_(2)Ⅲ_(2)Ⅳ_(2)structure suggests this megacomplex could act as an integral unit in electron transfer and proton pumping.The conformational changes from different states suggest a plausible regulatory mechanism for the MCⅠ_(2)Ⅲ_(2)Ⅳ_(2)activation/deactivation process.展开更多
Metal–organic frameworks(MOFs) are crystalline porous materials with tunable properties, exhibiting great potential in gas adsorption, separation and catalysis.[1,2]It is challenging to visualize MOFs with transmissi...Metal–organic frameworks(MOFs) are crystalline porous materials with tunable properties, exhibiting great potential in gas adsorption, separation and catalysis.[1,2]It is challenging to visualize MOFs with transmission electron microscopy(TEM) due to their inherent instability under electron beam irradiation. Here, we employ cryo-electron microscopy(cryoEM) to capture images of MOF ZIF-8, revealing inverted-space structural information at a resolution of up to about 1.7A and enhancing its critical electron dose to around 20 e^(-)/A^(2). In addition, it is confirmed by electron-beam irradiation experiments that the high voltage could effectively mitigate the radiolysis, and the structure of ZIF-8 is more stable along the [100] direction under electron beam irradiation. Meanwhile, since the high-resolution electron microscope images are modulated by contrast transfer function(CTF) and it is difficult to determine the positions corresponding to the atomic columns directly from the images. We employ image deconvolution to eliminate the impact of CTF and obtain the structural images of ZIF-8. As a result, the heavy atom Zn and the organic imidazole ring within the organic framework can be distinguished from structural images.展开更多
Leptin receptor(LepR)signaling plays an essential role in balancing food intake and energy expenditure.The architec-ture of LepR signaling assembly is critical for its function.In this study,we determined the structur...Leptin receptor(LepR)signaling plays an essential role in balancing food intake and energy expenditure.The architec-ture of LepR signaling assembly is critical for its function.In this study,we determined the structures of three distinct conformations of human leptin–LepR using cryo-electron microscopy at resolutions of 3.88,3.77,and 3.58Å.Both 2:2 and 3:3 stoichiometric assemblies were observed,and the complexes exhibited asymmetric open conformations.Lep-tin undergoes substantial rearrangement of its flexible regions to accommodate binding to LepR.The assembled leptin–LepR complexes connect through a“hand-in-hand”geometry.The open,interlocked 3:3 trimeric assembly results from the engagement of a third leptin–LepR heterodimer with a 2:2 dimer.The asymmetric geometry of LepR is substantially distinct from that of other gp130 cytokine homologs,and that may be due to the twisted and rigid interface between the D3 and D4 domains.These results highlight the distinct engagement of leptin with LepR and provide important insights into the structural plasticity of LepR-signaling assemblies.展开更多
Drug discovery is a crucial part of human healthcare and has dramatically benefited human lifespan and life quality in recent centuries, however, it is usually time-and effort-consuming. Structural biology has been de...Drug discovery is a crucial part of human healthcare and has dramatically benefited human lifespan and life quality in recent centuries, however, it is usually time-and effort-consuming. Structural biology has been demonstrated as a powerful tool to accelerate drug development. Among different techniques, cryo-electron microscopy(cryo-EM) is emerging as the mainstream of structure determination of biomacromolecules in the past decade and has received increasing attention from the pharmaceutical industry. Although cryo-EM still has limitations in resolution, speed and throughput, a growing number of innovative drugs are being developed with the help of cryo-EM. Here, we aim to provide an overview of how cryo-EM techniques are applied to facilitate drug discovery. The development and typical workflow of cryo-EM technique will be briefly introduced, followed by its specific applications in structure-based drug design, fragment-based drug discovery, proteolysis targeting chimeras, antibody drug development and drug repurposing. Besides cryo-EM, drug discovery innovation usually involves other state-of-the-art techniques such as artificial intelligence(AI), which is increasingly active in diverse areas. The combination of cryo-EM and AI provides an opportunity to minimize limitations of cryo-EM such as automation, throughput and interpretation of mediumresolution maps, and tends to be the new direction of future development of cryo-EM. The rapid development of cryo-EM will make it as an indispensable part of modern drug discovery.展开更多
Antibodies play critical roles in neutralizing viral infections and are increasingly used as therapeutic drugs and diagnostic tools. Structural studies on virus-antibody immune complexes are important for better under...Antibodies play critical roles in neutralizing viral infections and are increasingly used as therapeutic drugs and diagnostic tools. Structural studies on virus-antibody immune complexes are important for better understanding the molecular mechanisms of antibody-mediated neutralization and also provide valuable information for structure-based vaccine design.Cryo-electron microscopy(cryo-EM) has recently matured as a powerful structural technique for studying bio-macromolecular complexes. When combined with X-ray crystallography, cryo-EM provides a routine approach for structurally characterizing the immune complexes formed between icosahedral viruses and their antibodies. In this review, recent advances in the structural understanding of virus-antibody interactions are outlined for whole virions with icosahedral T = pseudo 3(picornaviruses) and T = 3(flaviviruses) architectures, focusing on the dynamic nature of viral shells in different functional states. Glycoprotein complexes from pleomorphic enveloped viruses are also discussed as immune complex antigens. Improving our understanding of viral epitope structures using virus-based platforms would provide a fundamental road map for future vaccine development.展开更多
As we know more about Zika virus(ZIKV), as well as its linkage to birth defects(microcephaly) and autoimmune neurological syndromes, we realize the importance of developing an efficient vaccine against it. Zika virus ...As we know more about Zika virus(ZIKV), as well as its linkage to birth defects(microcephaly) and autoimmune neurological syndromes, we realize the importance of developing an efficient vaccine against it. Zika virus disease has affected many countries and is becoming a major public health concern. To deal with the infection of ZIKV, plenty of experiments have been done on selection of neutralizing antibodies that can target the envelope(E) protein on the surface of the virion. However, the existence of antibody-dependent enhancement(ADE) effect might limit the use of them as therapeutic candidates. In this review, we classify the neutralizing antibodies against ZIKV based on the epitopes and summarize the resolved structural information on antibody/antigen complex from X-ray crystallography and cryo-electron microscopy(cryo-EM), which might be useful for further development of potent neutralizing antibodies and vaccines toward clinical use.展开更多
The human Shwachman-Diamond syndrome （SDS） is an autosomal recessive disease caused by mutations in a highly conserved ribosome assembly factor SBDS. The functional role of SBDS is to cooperate with another assembly...The human Shwachman-Diamond syndrome （SDS） is an autosomal recessive disease caused by mutations in a highly conserved ribosome assembly factor SBDS. The functional role of SBDS is to cooperate with another assembly factor, elongation factor l-like （Eft1）, to pro- mote the release of eukaryotic initiation factor 6 （elF6） from the late-stage cytoplasmic 60S precursors. In the present work, we characterized, both biochemically and structurally, the interaction between the 60S subunit and SBDS protein （Sdolp） from yeast. Our data show that Sdolp interacts tightly with the mature 60S subunit in vitro through its domain I and II, and is capable of bridging two 60S subunits to form a stable 2：2 dimer. Structural analysis indicates that Sdolp bind to the ribosomal P-site, in the proximity of uL16 and uL5, and with direct contact to H69 and H38. The dynamic nature of Sdolp on the 60S subunit, together with its strategic binding position, suggests a surveillance role of Sdolp in monitoring the conformational maturation of the ribosomal P-site. Altogether, our data support a con- formational signal-relay cascade during late-stage 60S maturation, involving uL16, Sdolp, and Efllp, which interrogates the functional P-site to control the departure of the anti-association factor elF6.展开更多
Recent technical breakthroughs in cryo-electron microscopy（cryo-EM） revolutionized structural biology, which led to the 2017 Nobel Prize in chemistry being awarded to three scientists, Jacques Dubochet, Joachim Fran...Recent technical breakthroughs in cryo-electron microscopy（cryo-EM） revolutionized structural biology, which led to the 2017 Nobel Prize in chemistry being awarded to three scientists, Jacques Dubochet, Joachim Frank, and Richard Henderson, who made groundbreaking contributions to the development of cryo-EM. In this review, I will give a comprehensive review of the developmental history of cryo-EM, the technical aspects of the breakthrough in cryo-EM leading to the structural biology revolution, including electron microscopy, image recording devices and image processing algorithms,and the major scientific achievements by Chinese researchers employing cryo-EM, covering protein complexes involved in or related to gene expression and regulation, protein synthesis and degradation, membrane proteins, immunity, and viruses.Finally, I will give a perspective outlook on the development of cryo-EM in the future.展开更多
基金supported by funds from the National Natural Science Foundation of China(32030056 and 32100962)the Tsinghua University Spring Breeze Fund(20201080572)+4 种基金the National Science Fund for Distinguished Young Scholars(3210110055)the China Postdoctoral Science Foundation(2020TQ0178,2020M680519,and 2020M680521)the Guangdong Basic and Applied Basic Research Foundation(2023B1515020039)the Shenzhen Science and Technology Program(RCYX20221008092904016)the Shenzhen University 2035 Program for Excellent Research(2022C012).
文摘Mammalian mitochondrial electron transport chain complexes are the most important and complicated protein machinery in mitochondria.Although this system has been studied for more than a century,its composition and molecular mechanism are still largely unknown.Here we report the high-resolution cryo-electron microscopy(Cryo-EM)structures of porcine respiratory chain megacomplex-Ⅰ_(2)Ⅲ_(2)Ⅳ_(2)(MCⅠ_(2)Ⅲ_(2)Ⅳ_(2))in five different conformations,including State 1,State 2,Mid 1,Mid 2,and Mid 3.High-resolution Cryo-EM imaging,combined with super-resolution gated stimulated emission depletion microscopy(gSTED),strongly supports the formation of MCⅠ_(2)Ⅲ_(2)Ⅳ_(2)in live cells.Each MCⅠ_(2)Ⅲ_(2)Ⅳ_(2)structure contains 141 subunits(70 different kinds of peptides,2.9 MDa)in total with 240 transmembrane helices.The mutual influence among CⅠ,CⅢ,and CⅣshown in the MCⅠ_(2)Ⅲ_(2)Ⅳ_(2)structure suggests this megacomplex could act as an integral unit in electron transfer and proton pumping.The conformational changes from different states suggest a plausible regulatory mechanism for the MCⅠ_(2)Ⅲ_(2)Ⅳ_(2)activation/deactivation process.
基金Project supported by the National Natural Science Foundation of China(Grant Nos.12074409 and 12374021)。
文摘Metal–organic frameworks(MOFs) are crystalline porous materials with tunable properties, exhibiting great potential in gas adsorption, separation and catalysis.[1,2]It is challenging to visualize MOFs with transmission electron microscopy(TEM) due to their inherent instability under electron beam irradiation. Here, we employ cryo-electron microscopy(cryoEM) to capture images of MOF ZIF-8, revealing inverted-space structural information at a resolution of up to about 1.7A and enhancing its critical electron dose to around 20 e^(-)/A^(2). In addition, it is confirmed by electron-beam irradiation experiments that the high voltage could effectively mitigate the radiolysis, and the structure of ZIF-8 is more stable along the [100] direction under electron beam irradiation. Meanwhile, since the high-resolution electron microscope images are modulated by contrast transfer function(CTF) and it is difficult to determine the positions corresponding to the atomic columns directly from the images. We employ image deconvolution to eliminate the impact of CTF and obtain the structural images of ZIF-8. As a result, the heavy atom Zn and the organic imidazole ring within the organic framework can be distinguished from structural images.
基金sup-ported by the National Key Research and Development Program of China (grant numbers 2020YFA0509202).
文摘Leptin receptor(LepR)signaling plays an essential role in balancing food intake and energy expenditure.The architec-ture of LepR signaling assembly is critical for its function.In this study,we determined the structures of three distinct conformations of human leptin–LepR using cryo-electron microscopy at resolutions of 3.88,3.77,and 3.58Å.Both 2:2 and 3:3 stoichiometric assemblies were observed,and the complexes exhibited asymmetric open conformations.Lep-tin undergoes substantial rearrangement of its flexible regions to accommodate binding to LepR.The assembled leptin–LepR complexes connect through a“hand-in-hand”geometry.The open,interlocked 3:3 trimeric assembly results from the engagement of a third leptin–LepR heterodimer with a 2:2 dimer.The asymmetric geometry of LepR is substantially distinct from that of other gp130 cytokine homologs,and that may be due to the twisted and rigid interface between the D3 and D4 domains.These results highlight the distinct engagement of leptin with LepR and provide important insights into the structural plasticity of LepR-signaling assemblies.
基金funded by the National Natural Science Foundation of China (NSFC, 31900046, 81972085, 82172465 and 32161133022)the Guangdong Provincial Key Laboratory of Advanced Biomaterials (2022B1212010003)+7 种基金the National Science and Technology Innovation 2030 Major Program (2022ZD0211900)the Shenzhen Key Laboratory of Computer Aided Drug Discovery (ZDSYS20201230165400001)the Chinese Academy of Science President’s International Fellowship Initiative (PIFI)(2020FSB0003)the Guangdong Retired Expert (granted by Guangdong Province)the Shenzhen Pengcheng ScientistNSFC-SNSF Funding (32161133022)Alpha Mol&SIAT Joint LaboratoryShenzhen Government Top-talent Working Funding and Guangdong Province Academician Work Funding。
文摘Drug discovery is a crucial part of human healthcare and has dramatically benefited human lifespan and life quality in recent centuries, however, it is usually time-and effort-consuming. Structural biology has been demonstrated as a powerful tool to accelerate drug development. Among different techniques, cryo-electron microscopy(cryo-EM) is emerging as the mainstream of structure determination of biomacromolecules in the past decade and has received increasing attention from the pharmaceutical industry. Although cryo-EM still has limitations in resolution, speed and throughput, a growing number of innovative drugs are being developed with the help of cryo-EM. Here, we aim to provide an overview of how cryo-EM techniques are applied to facilitate drug discovery. The development and typical workflow of cryo-EM technique will be briefly introduced, followed by its specific applications in structure-based drug design, fragment-based drug discovery, proteolysis targeting chimeras, antibody drug development and drug repurposing. Besides cryo-EM, drug discovery innovation usually involves other state-of-the-art techniques such as artificial intelligence(AI), which is increasingly active in diverse areas. The combination of cryo-EM and AI provides an opportunity to minimize limitations of cryo-EM such as automation, throughput and interpretation of mediumresolution maps, and tends to be the new direction of future development of cryo-EM. The rapid development of cryo-EM will make it as an indispensable part of modern drug discovery.
基金the National Natural Science Foundation of China(Grant Nos.31570161 and 31770169)the“One-Three-Five”Strategic Programs of the Wuhan Institute of Virology,Chinese Academy of Sciences(Grant No.Y605211SA3).
文摘Antibodies play critical roles in neutralizing viral infections and are increasingly used as therapeutic drugs and diagnostic tools. Structural studies on virus-antibody immune complexes are important for better understanding the molecular mechanisms of antibody-mediated neutralization and also provide valuable information for structure-based vaccine design.Cryo-electron microscopy(cryo-EM) has recently matured as a powerful structural technique for studying bio-macromolecular complexes. When combined with X-ray crystallography, cryo-EM provides a routine approach for structurally characterizing the immune complexes formed between icosahedral viruses and their antibodies. In this review, recent advances in the structural understanding of virus-antibody interactions are outlined for whole virions with icosahedral T = pseudo 3(picornaviruses) and T = 3(flaviviruses) architectures, focusing on the dynamic nature of viral shells in different functional states. Glycoprotein complexes from pleomorphic enveloped viruses are also discussed as immune complex antigens. Improving our understanding of viral epitope structures using virus-based platforms would provide a fundamental road map for future vaccine development.
基金funded by the External Cooperation Program of Chinese Academy of Sciences (Grant No. 153211KYSB20160001)the National Key Research and Development Program of China (Grant No. 2016YFC1202902)+1 种基金the Key Program of Chinese Academy of Sciences (Grant No. ZDRW-ZS2016-4)funded by FNLCR Contract HHSN261200800001E
文摘As we know more about Zika virus(ZIKV), as well as its linkage to birth defects(microcephaly) and autoimmune neurological syndromes, we realize the importance of developing an efficient vaccine against it. Zika virus disease has affected many countries and is becoming a major public health concern. To deal with the infection of ZIKV, plenty of experiments have been done on selection of neutralizing antibodies that can target the envelope(E) protein on the surface of the virion. However, the existence of antibody-dependent enhancement(ADE) effect might limit the use of them as therapeutic candidates. In this review, we classify the neutralizing antibodies against ZIKV based on the epitopes and summarize the resolved structural information on antibody/antigen complex from X-ray crystallography and cryo-electron microscopy(cryo-EM), which might be useful for further development of potent neutralizing antibodies and vaccines toward clinical use.
文摘The human Shwachman-Diamond syndrome （SDS） is an autosomal recessive disease caused by mutations in a highly conserved ribosome assembly factor SBDS. The functional role of SBDS is to cooperate with another assembly factor, elongation factor l-like （Eft1）, to pro- mote the release of eukaryotic initiation factor 6 （elF6） from the late-stage cytoplasmic 60S precursors. In the present work, we characterized, both biochemically and structurally, the interaction between the 60S subunit and SBDS protein （Sdolp） from yeast. Our data show that Sdolp interacts tightly with the mature 60S subunit in vitro through its domain I and II, and is capable of bridging two 60S subunits to form a stable 2：2 dimer. Structural analysis indicates that Sdolp bind to the ribosomal P-site, in the proximity of uL16 and uL5, and with direct contact to H69 and H38. The dynamic nature of Sdolp on the 60S subunit, together with its strategic binding position, suggests a surveillance role of Sdolp in monitoring the conformational maturation of the ribosomal P-site. Altogether, our data support a con- formational signal-relay cascade during late-stage 60S maturation, involving uL16, Sdolp, and Efllp, which interrogates the functional P-site to control the departure of the anti-association factor elF6.
基金Project supported by the National Key Research and Development Program of China(Grant No.2017YFA0504700)the National Natural Science Foundation of China(Grant Nos.31570732 and 31770785)
文摘Recent technical breakthroughs in cryo-electron microscopy（cryo-EM） revolutionized structural biology, which led to the 2017 Nobel Prize in chemistry being awarded to three scientists, Jacques Dubochet, Joachim Frank, and Richard Henderson, who made groundbreaking contributions to the development of cryo-EM. In this review, I will give a comprehensive review of the developmental history of cryo-EM, the technical aspects of the breakthrough in cryo-EM leading to the structural biology revolution, including electron microscopy, image recording devices and image processing algorithms,and the major scientific achievements by Chinese researchers employing cryo-EM, covering protein complexes involved in or related to gene expression and regulation, protein synthesis and degradation, membrane proteins, immunity, and viruses.Finally, I will give a perspective outlook on the development of cryo-EM in the future.
