The Experts below are selected from a list of 360 Experts worldwide ranked by ideXlab platform
Marc Messerschmidt - One of the best experts on this subject based on the ideXlab platform.
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segmented flow generator for serial Crystallography at the european x ray free electron laser
Nature Communications, 2020Co-Authors: Austin Echelmeier, Sabine Botha, Marc Messerschmidt, Jorvani Cruz Villarreal, Daihyun Kim, Jesse Coe, Darren Thifault, Ana Egatzgomez, Sahir GandhiAbstract:Serial femtosecond Crystallography (SFX) with X-ray free electron lasers (XFELs) allows structure determination of membrane Proteins and time-resolved Crystallography. Common liquid sample delivery continuously jets the Protein Crystal suspension into the path of the XFEL, wasting a vast amount of sample due to the pulsed nature of all current XFEL sources. The European XFEL (EuXFEL) delivers femtosecond (fs) X-ray pulses in trains spaced 100 ms apart whereas pulses within trains are currently separated by 889 ns. Therefore, continuous sample delivery via fast jets wastes >99% of sample. Here, we introduce a microfluidic device delivering Crystal laden droplets segmented with an immiscible oil reducing sample waste and demonstrate droplet injection at the EuXFEL compatible with high pressure liquid delivery of an SFX experiment. While achieving ~60% reduction in sample waste, we determine the structure of the enzyme 3-deoxy-D-manno-octulosonate-8-phosphate synthase from microCrystals delivered in droplets revealing distinct structural features not previously reported. Due to the pulsed nature of X-ray free electron laser (XFEL) instruments the majority of Protein Crystals, which are injected using continuous jet injection techniques are wasted. Here, the authors present a microfluidic device to deliver aqueous Protein Crystal laden droplets segmented with an immiscible oil and demonstrate that with this device an approx. 60% reduction in sample waste was achieved for data collection of 3-deoxy-D-manno-octulosonate 8-phosphate synthase Crystals at the EuXFEL.
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de novo Protein Crystal structure determination from x ray free electron laser data
Nature, 2014Co-Authors: Thomas R M Barends, Lutz Foucar, Sabine Botha, Bruce R Doak, Robert L Shoeman, Karol Nass, Jason E Koglin, Garth J Williams, Sebastien Boutet, Marc MesserschmidtAbstract:The determination of Protein Crystal structures is hampered by the need for macroscopic Crystals. X-ray free-electron lasers (FELs) provide extremely intense pulses of femtosecond duration, which allow data collection from nanometre- to micrometre-sized Crystals in a 'diffraction-before-destruction' approach. So far, all Protein structure determinations carried out using FELs have been based on previous knowledge of related, known structures. Here we show that X-ray FEL data can be used for de novo Protein structure determination, that is, without previous knowledge about the structure. Using the emerging technique of serial femtosecond Crystallography, we performed single-wavelength anomalous scattering measurements on microCrystals of the well-established model system lysozyme, in complex with a lanthanide compound. Using Monte-Carlo integration, we obtained high-quality diffraction intensities from which experimental phases could be determined, resulting in an experimental electron density map good enough for automated building of the Protein structure. This demonstrates the feasibility of determining novel Protein structures using FELs. We anticipate that serial femtosecond Crystallography will become an important tool for the structure determination of Proteins that are difficult to Crystallize, such as membrane Proteins.
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de novo Protein Crystal structure determination from x ray free electron laser data
Nature, 2014Co-Authors: Thomas R M Barends, Lutz Foucar, Sabine Botha, Bruce R Doak, Robert L Shoeman, Karol Nass, Jason E Koglin, Garth J Williams, Sebastien Boutet, Marc MesserschmidtAbstract:Femtosecond Crystallography with an X-ray free-electron laser is used to analyse micrometre-sized Protein Crystals, generating a high-resolution structure of the Protein without previous knowledge of what it looks like. X-ray Crystallographers typically spend a great deal of time optimizing Crystallization conditions to obtain the large, well-ordered Crystals needed to generate high-quality data sets. It was shown recently that extremely short and intense pulses of X-rays from X-ray free-electron lasers can be used to obtain diffraction data on nano-to-micrometre-sized Protein Crystals before radiation damage to the Crystal occurs. The hope is that this approach — called serial femtosecond Crystallography — will produce structures of Proteins and Protein complexes that do not yield macroscopic, well-ordered Crystals. One of the major limitations of serial femtosecond Crystallography is that it has not been possible to solve the structure of a Protein without prior knowledge of related, known structures. In this paper, the authors show how serial femtosecond Crystallography with an X-ray free-electron laser can be used to experimentally solve the 'phase problem', generating a high-resolution structure of a Protein without a priori knowledge of what the Protein looks like. The determination of Protein Crystal structures is hampered by the need for macroscopic Crystals. X-ray free-electron lasers (FELs) provide extremely intense pulses of femtosecond duration, which allow data collection from nanometre- to micrometre-sized Crystals1,2,3,4 in a ‘diffraction-before-destruction’ approach. So far, all Protein structure determinations carried out using FELs have been based on previous knowledge of related, known structures1,2,3,4,5. Here we show that X-ray FEL data can be used for de novo Protein structure determination, that is, without previous knowledge about the structure. Using the emerging technique of serial femtosecond Crystallography1,2,3,4,6, we performed single-wavelength anomalous scattering measurements on microCrystals of the well-established model system lysozyme, in complex with a lanthanide compound. Using Monte-Carlo integration6,7, we obtained high-quality diffraction intensities from which experimental phases could be determined, resulting in an experimental electron density map good enough for automated building of the Protein structure. This demonstrates the feasibility of determining novel Protein structures using FELs. We anticipate that serial femtosecond Crystallography will become an important tool for the structure determination of Proteins that are difficult to Crystallize, such as membrane Proteins1,2,8.
Sabine Botha - One of the best experts on this subject based on the ideXlab platform.
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segmented flow generator for serial Crystallography at the european x ray free electron laser
Nature Communications, 2020Co-Authors: Austin Echelmeier, Sabine Botha, Marc Messerschmidt, Jorvani Cruz Villarreal, Daihyun Kim, Jesse Coe, Darren Thifault, Ana Egatzgomez, Sahir GandhiAbstract:Serial femtosecond Crystallography (SFX) with X-ray free electron lasers (XFELs) allows structure determination of membrane Proteins and time-resolved Crystallography. Common liquid sample delivery continuously jets the Protein Crystal suspension into the path of the XFEL, wasting a vast amount of sample due to the pulsed nature of all current XFEL sources. The European XFEL (EuXFEL) delivers femtosecond (fs) X-ray pulses in trains spaced 100 ms apart whereas pulses within trains are currently separated by 889 ns. Therefore, continuous sample delivery via fast jets wastes >99% of sample. Here, we introduce a microfluidic device delivering Crystal laden droplets segmented with an immiscible oil reducing sample waste and demonstrate droplet injection at the EuXFEL compatible with high pressure liquid delivery of an SFX experiment. While achieving ~60% reduction in sample waste, we determine the structure of the enzyme 3-deoxy-D-manno-octulosonate-8-phosphate synthase from microCrystals delivered in droplets revealing distinct structural features not previously reported. Due to the pulsed nature of X-ray free electron laser (XFEL) instruments the majority of Protein Crystals, which are injected using continuous jet injection techniques are wasted. Here, the authors present a microfluidic device to deliver aqueous Protein Crystal laden droplets segmented with an immiscible oil and demonstrate that with this device an approx. 60% reduction in sample waste was achieved for data collection of 3-deoxy-D-manno-octulosonate 8-phosphate synthase Crystals at the EuXFEL.
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de novo Protein Crystal structure determination from x ray free electron laser data
Nature, 2014Co-Authors: Thomas R M Barends, Lutz Foucar, Sabine Botha, Bruce R Doak, Robert L Shoeman, Karol Nass, Jason E Koglin, Garth J Williams, Sebastien Boutet, Marc MesserschmidtAbstract:The determination of Protein Crystal structures is hampered by the need for macroscopic Crystals. X-ray free-electron lasers (FELs) provide extremely intense pulses of femtosecond duration, which allow data collection from nanometre- to micrometre-sized Crystals in a 'diffraction-before-destruction' approach. So far, all Protein structure determinations carried out using FELs have been based on previous knowledge of related, known structures. Here we show that X-ray FEL data can be used for de novo Protein structure determination, that is, without previous knowledge about the structure. Using the emerging technique of serial femtosecond Crystallography, we performed single-wavelength anomalous scattering measurements on microCrystals of the well-established model system lysozyme, in complex with a lanthanide compound. Using Monte-Carlo integration, we obtained high-quality diffraction intensities from which experimental phases could be determined, resulting in an experimental electron density map good enough for automated building of the Protein structure. This demonstrates the feasibility of determining novel Protein structures using FELs. We anticipate that serial femtosecond Crystallography will become an important tool for the structure determination of Proteins that are difficult to Crystallize, such as membrane Proteins.
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de novo Protein Crystal structure determination from x ray free electron laser data
Nature, 2014Co-Authors: Thomas R M Barends, Lutz Foucar, Sabine Botha, Bruce R Doak, Robert L Shoeman, Karol Nass, Jason E Koglin, Garth J Williams, Sebastien Boutet, Marc MesserschmidtAbstract:Femtosecond Crystallography with an X-ray free-electron laser is used to analyse micrometre-sized Protein Crystals, generating a high-resolution structure of the Protein without previous knowledge of what it looks like. X-ray Crystallographers typically spend a great deal of time optimizing Crystallization conditions to obtain the large, well-ordered Crystals needed to generate high-quality data sets. It was shown recently that extremely short and intense pulses of X-rays from X-ray free-electron lasers can be used to obtain diffraction data on nano-to-micrometre-sized Protein Crystals before radiation damage to the Crystal occurs. The hope is that this approach — called serial femtosecond Crystallography — will produce structures of Proteins and Protein complexes that do not yield macroscopic, well-ordered Crystals. One of the major limitations of serial femtosecond Crystallography is that it has not been possible to solve the structure of a Protein without prior knowledge of related, known structures. In this paper, the authors show how serial femtosecond Crystallography with an X-ray free-electron laser can be used to experimentally solve the 'phase problem', generating a high-resolution structure of a Protein without a priori knowledge of what the Protein looks like. The determination of Protein Crystal structures is hampered by the need for macroscopic Crystals. X-ray free-electron lasers (FELs) provide extremely intense pulses of femtosecond duration, which allow data collection from nanometre- to micrometre-sized Crystals1,2,3,4 in a ‘diffraction-before-destruction’ approach. So far, all Protein structure determinations carried out using FELs have been based on previous knowledge of related, known structures1,2,3,4,5. Here we show that X-ray FEL data can be used for de novo Protein structure determination, that is, without previous knowledge about the structure. Using the emerging technique of serial femtosecond Crystallography1,2,3,4,6, we performed single-wavelength anomalous scattering measurements on microCrystals of the well-established model system lysozyme, in complex with a lanthanide compound. Using Monte-Carlo integration6,7, we obtained high-quality diffraction intensities from which experimental phases could be determined, resulting in an experimental electron density map good enough for automated building of the Protein structure. This demonstrates the feasibility of determining novel Protein structures using FELs. We anticipate that serial femtosecond Crystallography will become an important tool for the structure determination of Proteins that are difficult to Crystallize, such as membrane Proteins1,2,8.
Thomas R M Barends - One of the best experts on this subject based on the ideXlab platform.
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de novo Protein Crystal structure determination from x ray free electron laser data
Nature, 2014Co-Authors: Thomas R M Barends, Lutz Foucar, Sabine Botha, Bruce R Doak, Robert L Shoeman, Karol Nass, Jason E Koglin, Garth J Williams, Sebastien Boutet, Marc MesserschmidtAbstract:The determination of Protein Crystal structures is hampered by the need for macroscopic Crystals. X-ray free-electron lasers (FELs) provide extremely intense pulses of femtosecond duration, which allow data collection from nanometre- to micrometre-sized Crystals in a 'diffraction-before-destruction' approach. So far, all Protein structure determinations carried out using FELs have been based on previous knowledge of related, known structures. Here we show that X-ray FEL data can be used for de novo Protein structure determination, that is, without previous knowledge about the structure. Using the emerging technique of serial femtosecond Crystallography, we performed single-wavelength anomalous scattering measurements on microCrystals of the well-established model system lysozyme, in complex with a lanthanide compound. Using Monte-Carlo integration, we obtained high-quality diffraction intensities from which experimental phases could be determined, resulting in an experimental electron density map good enough for automated building of the Protein structure. This demonstrates the feasibility of determining novel Protein structures using FELs. We anticipate that serial femtosecond Crystallography will become an important tool for the structure determination of Proteins that are difficult to Crystallize, such as membrane Proteins.
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de novo Protein Crystal structure determination from x ray free electron laser data
Nature, 2014Co-Authors: Thomas R M Barends, Lutz Foucar, Sabine Botha, Bruce R Doak, Robert L Shoeman, Karol Nass, Jason E Koglin, Garth J Williams, Sebastien Boutet, Marc MesserschmidtAbstract:Femtosecond Crystallography with an X-ray free-electron laser is used to analyse micrometre-sized Protein Crystals, generating a high-resolution structure of the Protein without previous knowledge of what it looks like. X-ray Crystallographers typically spend a great deal of time optimizing Crystallization conditions to obtain the large, well-ordered Crystals needed to generate high-quality data sets. It was shown recently that extremely short and intense pulses of X-rays from X-ray free-electron lasers can be used to obtain diffraction data on nano-to-micrometre-sized Protein Crystals before radiation damage to the Crystal occurs. The hope is that this approach — called serial femtosecond Crystallography — will produce structures of Proteins and Protein complexes that do not yield macroscopic, well-ordered Crystals. One of the major limitations of serial femtosecond Crystallography is that it has not been possible to solve the structure of a Protein without prior knowledge of related, known structures. In this paper, the authors show how serial femtosecond Crystallography with an X-ray free-electron laser can be used to experimentally solve the 'phase problem', generating a high-resolution structure of a Protein without a priori knowledge of what the Protein looks like. The determination of Protein Crystal structures is hampered by the need for macroscopic Crystals. X-ray free-electron lasers (FELs) provide extremely intense pulses of femtosecond duration, which allow data collection from nanometre- to micrometre-sized Crystals1,2,3,4 in a ‘diffraction-before-destruction’ approach. So far, all Protein structure determinations carried out using FELs have been based on previous knowledge of related, known structures1,2,3,4,5. Here we show that X-ray FEL data can be used for de novo Protein structure determination, that is, without previous knowledge about the structure. Using the emerging technique of serial femtosecond Crystallography1,2,3,4,6, we performed single-wavelength anomalous scattering measurements on microCrystals of the well-established model system lysozyme, in complex with a lanthanide compound. Using Monte-Carlo integration6,7, we obtained high-quality diffraction intensities from which experimental phases could be determined, resulting in an experimental electron density map good enough for automated building of the Protein structure. This demonstrates the feasibility of determining novel Protein structures using FELs. We anticipate that serial femtosecond Crystallography will become an important tool for the structure determination of Proteins that are difficult to Crystallize, such as membrane Proteins1,2,8.
Karol Nass - One of the best experts on this subject based on the ideXlab platform.
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de novo Protein Crystal structure determination from x ray free electron laser data
Nature, 2014Co-Authors: Thomas R M Barends, Lutz Foucar, Sabine Botha, Bruce R Doak, Robert L Shoeman, Karol Nass, Jason E Koglin, Garth J Williams, Sebastien Boutet, Marc MesserschmidtAbstract:The determination of Protein Crystal structures is hampered by the need for macroscopic Crystals. X-ray free-electron lasers (FELs) provide extremely intense pulses of femtosecond duration, which allow data collection from nanometre- to micrometre-sized Crystals in a 'diffraction-before-destruction' approach. So far, all Protein structure determinations carried out using FELs have been based on previous knowledge of related, known structures. Here we show that X-ray FEL data can be used for de novo Protein structure determination, that is, without previous knowledge about the structure. Using the emerging technique of serial femtosecond Crystallography, we performed single-wavelength anomalous scattering measurements on microCrystals of the well-established model system lysozyme, in complex with a lanthanide compound. Using Monte-Carlo integration, we obtained high-quality diffraction intensities from which experimental phases could be determined, resulting in an experimental electron density map good enough for automated building of the Protein structure. This demonstrates the feasibility of determining novel Protein structures using FELs. We anticipate that serial femtosecond Crystallography will become an important tool for the structure determination of Proteins that are difficult to Crystallize, such as membrane Proteins.
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de novo Protein Crystal structure determination from x ray free electron laser data
Nature, 2014Co-Authors: Thomas R M Barends, Lutz Foucar, Sabine Botha, Bruce R Doak, Robert L Shoeman, Karol Nass, Jason E Koglin, Garth J Williams, Sebastien Boutet, Marc MesserschmidtAbstract:Femtosecond Crystallography with an X-ray free-electron laser is used to analyse micrometre-sized Protein Crystals, generating a high-resolution structure of the Protein without previous knowledge of what it looks like. X-ray Crystallographers typically spend a great deal of time optimizing Crystallization conditions to obtain the large, well-ordered Crystals needed to generate high-quality data sets. It was shown recently that extremely short and intense pulses of X-rays from X-ray free-electron lasers can be used to obtain diffraction data on nano-to-micrometre-sized Protein Crystals before radiation damage to the Crystal occurs. The hope is that this approach — called serial femtosecond Crystallography — will produce structures of Proteins and Protein complexes that do not yield macroscopic, well-ordered Crystals. One of the major limitations of serial femtosecond Crystallography is that it has not been possible to solve the structure of a Protein without prior knowledge of related, known structures. In this paper, the authors show how serial femtosecond Crystallography with an X-ray free-electron laser can be used to experimentally solve the 'phase problem', generating a high-resolution structure of a Protein without a priori knowledge of what the Protein looks like. The determination of Protein Crystal structures is hampered by the need for macroscopic Crystals. X-ray free-electron lasers (FELs) provide extremely intense pulses of femtosecond duration, which allow data collection from nanometre- to micrometre-sized Crystals1,2,3,4 in a ‘diffraction-before-destruction’ approach. So far, all Protein structure determinations carried out using FELs have been based on previous knowledge of related, known structures1,2,3,4,5. Here we show that X-ray FEL data can be used for de novo Protein structure determination, that is, without previous knowledge about the structure. Using the emerging technique of serial femtosecond Crystallography1,2,3,4,6, we performed single-wavelength anomalous scattering measurements on microCrystals of the well-established model system lysozyme, in complex with a lanthanide compound. Using Monte-Carlo integration6,7, we obtained high-quality diffraction intensities from which experimental phases could be determined, resulting in an experimental electron density map good enough for automated building of the Protein structure. This demonstrates the feasibility of determining novel Protein structures using FELs. We anticipate that serial femtosecond Crystallography will become an important tool for the structure determination of Proteins that are difficult to Crystallize, such as membrane Proteins1,2,8.
Sebastien Boutet - One of the best experts on this subject based on the ideXlab platform.
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de novo Protein Crystal structure determination from x ray free electron laser data
Nature, 2014Co-Authors: Thomas R M Barends, Lutz Foucar, Sabine Botha, Bruce R Doak, Robert L Shoeman, Karol Nass, Jason E Koglin, Garth J Williams, Sebastien Boutet, Marc MesserschmidtAbstract:The determination of Protein Crystal structures is hampered by the need for macroscopic Crystals. X-ray free-electron lasers (FELs) provide extremely intense pulses of femtosecond duration, which allow data collection from nanometre- to micrometre-sized Crystals in a 'diffraction-before-destruction' approach. So far, all Protein structure determinations carried out using FELs have been based on previous knowledge of related, known structures. Here we show that X-ray FEL data can be used for de novo Protein structure determination, that is, without previous knowledge about the structure. Using the emerging technique of serial femtosecond Crystallography, we performed single-wavelength anomalous scattering measurements on microCrystals of the well-established model system lysozyme, in complex with a lanthanide compound. Using Monte-Carlo integration, we obtained high-quality diffraction intensities from which experimental phases could be determined, resulting in an experimental electron density map good enough for automated building of the Protein structure. This demonstrates the feasibility of determining novel Protein structures using FELs. We anticipate that serial femtosecond Crystallography will become an important tool for the structure determination of Proteins that are difficult to Crystallize, such as membrane Proteins.
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de novo Protein Crystal structure determination from x ray free electron laser data
Nature, 2014Co-Authors: Thomas R M Barends, Lutz Foucar, Sabine Botha, Bruce R Doak, Robert L Shoeman, Karol Nass, Jason E Koglin, Garth J Williams, Sebastien Boutet, Marc MesserschmidtAbstract:Femtosecond Crystallography with an X-ray free-electron laser is used to analyse micrometre-sized Protein Crystals, generating a high-resolution structure of the Protein without previous knowledge of what it looks like. X-ray Crystallographers typically spend a great deal of time optimizing Crystallization conditions to obtain the large, well-ordered Crystals needed to generate high-quality data sets. It was shown recently that extremely short and intense pulses of X-rays from X-ray free-electron lasers can be used to obtain diffraction data on nano-to-micrometre-sized Protein Crystals before radiation damage to the Crystal occurs. The hope is that this approach — called serial femtosecond Crystallography — will produce structures of Proteins and Protein complexes that do not yield macroscopic, well-ordered Crystals. One of the major limitations of serial femtosecond Crystallography is that it has not been possible to solve the structure of a Protein without prior knowledge of related, known structures. In this paper, the authors show how serial femtosecond Crystallography with an X-ray free-electron laser can be used to experimentally solve the 'phase problem', generating a high-resolution structure of a Protein without a priori knowledge of what the Protein looks like. The determination of Protein Crystal structures is hampered by the need for macroscopic Crystals. X-ray free-electron lasers (FELs) provide extremely intense pulses of femtosecond duration, which allow data collection from nanometre- to micrometre-sized Crystals1,2,3,4 in a ‘diffraction-before-destruction’ approach. So far, all Protein structure determinations carried out using FELs have been based on previous knowledge of related, known structures1,2,3,4,5. Here we show that X-ray FEL data can be used for de novo Protein structure determination, that is, without previous knowledge about the structure. Using the emerging technique of serial femtosecond Crystallography1,2,3,4,6, we performed single-wavelength anomalous scattering measurements on microCrystals of the well-established model system lysozyme, in complex with a lanthanide compound. Using Monte-Carlo integration6,7, we obtained high-quality diffraction intensities from which experimental phases could be determined, resulting in an experimental electron density map good enough for automated building of the Protein structure. This demonstrates the feasibility of determining novel Protein structures using FELs. We anticipate that serial femtosecond Crystallography will become an important tool for the structure determination of Proteins that are difficult to Crystallize, such as membrane Proteins1,2,8.
