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285 results on '"Fromme, Raimund"'

1. Detection of a Geminate Photoproduct of Bovine Cytochrome c Oxidase by Time-Resolved Serial Femtosecond Crystallography

Author

Ishigami, Izumi, Carbajo, Sergio, Zatsepin, Nadia, Hikita, Masahide, Conrad, Chelsie E, Nelson, Garrett, Coe, Jesse, Basu, Shibom, Grant, Thomas, Seaberg, Matthew H, Sierra, Raymond G, Hunter, Mark S, Fromme, Petra, Fromme, Raimund, Rousseau, Denis L, and Yeh, Syun-Ru

Subjects
Inorganic Chemistry, Chemical Sciences, Acquired Cognitive Impairment, Brain Disorders, Cattle, Animals, Electron Transport Complex IV, Oxidation-Reduction, Copper, Ligands, Oxygen, Crystallography, X-Ray, Iron, Water, General Chemistry, Chemical sciences, Engineering
Abstract

Cytochrome c oxidase (CcO) is a large membrane-bound hemeprotein that catalyzes the reduction of dioxygen to water. Unlike classical dioxygen binding hemeproteins with a heme b group in their active sites, CcO has a unique binuclear center (BNC) composed of a copper atom (CuB) and a heme a3 iron, where O2 binds and is reduced to water. CO is a versatile O2 surrogate in ligand binding and escape reactions. Previous time-resolved spectroscopic studies of the CO complexes of bovine CcO (bCcO) revealed that photolyzing CO from the heme a3 iron leads to a metastable intermediate (CuB-CO), where CO is bound to CuB, before it escapes out of the BNC. Here, with a pump-probe based time-resolved serial femtosecond X-ray crystallography, we detected a geminate photoproduct of the bCcO-CO complex, where CO is dissociated from the heme a3 iron and moved to a temporary binding site midway between the CuB and the heme a3 iron, while the locations of the two metal centers and the conformation of Helix-X, housing the proximal histidine ligand of the heme a3 iron, remain in the CO complex state. This new structure, combined with other reported structures of bCcO, allows for a clearer definition of the ligand dissociation trajectory as well as the associated protein dynamics.

Published
2023

2. Room-temperature structural studies of SARS-CoV-2 protein NendoU with an X-ray free-electron laser.

Author

Jernigan, Rebecca J, Logeswaran, Dhenugen, Doppler, Diandra, Nagaratnam, Nirupa, Sonker, Mukul, Yang, Jay-How, Ketawala, Gihan, Martin-Garcia, Jose M, Shelby, Megan L, Grant, Thomas D, Mariani, Valerio, Tolstikova, Alexandra, Sheikh, Michelle Z, Yung, Mimi Cho, Coleman, Matthew A, Zaare, Sahba, Kaschner, Emily K, Rabbani, Mohammad Towshif, Nazari, Reza, Zacks, Michele A, Hayes, Brandon, Sierra, Raymond G, Hunter, Mark S, Lisova, Stella, Batyuk, Alexander, Kupitz, Christopher, Boutet, Sebastien, Hansen, Debra T, Kirian, Richard A, Schmidt, Marius, Fromme, Raimund, Frank, Matthias, Ros, Alexandra, Chen, Julian J-L, Botha, Sabine, and Fromme, Petra

Subjects
Humans, Crystallography, X-Ray, Lasers, Temperature, Electrons, COVID-19, SARS-CoV-2, NSP15, NendoU, X-ray free-electron laser, serial femtosecond crystallography, Prevention, Infectious Diseases, Emerging Infectious Diseases, Vaccine Related, Biodefense, Infection, Chemical Sciences, Biological Sciences, Information and Computing Sciences, Biophysics
Abstract

NendoU from SARS-CoV-2 is responsible for the virus's ability to evade the innate immune system by cleaving the polyuridine leader sequence of antisense viral RNA. Here we report the room-temperature structure of NendoU, solved by serial femtosecond crystallography at an X-ray free-electron laser to 2.6 Å resolution. The room-temperature structure provides insight into the flexibility, dynamics, and other intrinsic properties of NendoU, with indications that the enzyme functions as an allosteric switch. Functional studies examining cleavage specificity in solution and in crystals support the uridine-purine cleavage preference, and we demonstrate that enzyme activity is fully maintained in crystal form. Optimizing the purification of NendoU and identifying suitable crystallization conditions set the benchmark for future time-resolved serial femtosecond crystallography studies. This could advance the design of antivirals with higher efficacy in treating coronaviral infections, since drugs that block allosteric conformational changes are less prone to drug resistance.

Published
2023

3. Co-flow injection for serial crystallography at X-ray free-electron lasers.

Author

Doppler, Diandra, Rabbani, Mohammad, Letrun, Romain, Cruz Villarreal, Jorvani, Kim, Dai, Gandhi, Sahir, Egatz-Gomez, Ana, Sonker, Mukul, Chen, Joe, Koua, Faisal, Yang, Jayhow, Youssef, Mohamed, Mazalova, Victoria, Bajt, Saša, Shelby, Megan, Coleman, Matt, Wiedorn, Max, Knoska, Juraj, Schön, Silvan, Sato, Tokushi, Hunter, Mark, Hosseinizadeh, Ahmad, Kuptiz, Christopher, Nazari, Reza, Alvarez, Roberto, Karpos, Konstantinos, Zaare, Sahba, Dobson, Zachary, Discianno, Erin, Zhang, Shangji, Zook, James, Bielecki, Johan, de Wijn, Raphael, Round, Adam, Vagovic, Patrik, Kloos, Marco, Vakili, Mohammad, Ketawala, Gihan, Stander, Natasha, Olson, Tien, Morin, Katherine, Mondal, Jyotirmory, Nguyen, Jonathan, Meza-Aguilar, José, Kodis, Gerdenis, Vaiana, Sara, Martin-Garcia, Jose, Mariani, Valerio, Schwander, Peter, Schmidt, Marius, Messerschmidt, Marc, Ourmazd, Abbas, Zatsepin, Nadia, Weierstall, Uwe, Bruce, Barry, Mancuso, Adrian, Grant, Thomas, Barty, Anton, Chapman, Henry, Fromme, Raimund, Spence, John, Botha, Sabine, Fromme, Petra, Kirian, Richard, Ros, Alexandra, and Frank, Matthias

Subjects
3D printing, X-ray free-electron lasers, XFELs, microfluidic devices, sample consumption, serial crystallography, viscous media
Abstract

Serial femtosecond crystallography (SFX) is a powerful technique that exploits X-ray free-electron lasers to determine the structure of macro-molecules at room temperature. Despite the impressive exposition of structural details with this novel crystallographic approach, the methods currently available to introduce crystals into the path of the X-ray beam sometimes exhibit serious drawbacks. Samples requiring liquid injection of crystal slurries consume large quantities of crystals (at times up to a gram of protein per data set), may not be compatible with vacuum configurations on beamlines or provide a high background due to additional sheathing liquids present during the injection. Proposed and characterized here is the use of an immiscible inert oil phase to supplement the flow of sample in a hybrid microfluidic 3D-printed co-flow device. Co-flow generation is reported with sample and oil phases flowing in parallel, resulting in stable injection conditions for two different resin materials experimentally. A numerical model is presented that adequately predicts these flow-rate conditions. The co-flow generating devices reduce crystal clogging effects, have the potential to conserve protein crystal samples up to 95% and will allow degradation-free light-induced time-resolved SFX.

Published
2022

4. Time-resolved serial femtosecond crystallography at the European XFEL

Author

Pandey, Suraj, Bean, Richard, Sato, Tokushi, Poudyal, Ishwor, Bielecki, Johan, Cruz Villarreal, Jorvani, Yefanov, Oleksandr, Mariani, Valerio, White, Thomas A, Kupitz, Christopher, Hunter, Mark, Abdellatif, Mohamed H, Bajt, Saša, Bondar, Valerii, Echelmeier, Austin, Doppler, Diandra, Emons, Moritz, Frank, Matthias, Fromme, Raimund, Gevorkov, Yaroslav, Giovanetti, Gabriele, Jiang, Man, Kim, Daihyun, Kim, Yoonhee, Kirkwood, Henry, Klimovskaia, Anna, Knoska, Juraj, Koua, Faisal HM, Letrun, Romain, Lisova, Stella, Maia, Luis, Mazalova, Victoria, Meza, Domingo, Michelat, Thomas, Ourmazd, Abbas, Palmer, Guido, Ramilli, Marco, Schubert, Robin, Schwander, Peter, Silenzi, Alessandro, Sztuk-Dambietz, Jolanta, Tolstikova, Alexandra, Chapman, Henry N, Ros, Alexandra, Barty, Anton, Fromme, Petra, Mancuso, Adrian P, and Schmidt, Marius

Subjects
Biochemistry and Cell Biology, Biological Sciences, Bacterial Proteins, Crystallography, X-Ray, Light, Models, Molecular, Photoreceptors, Microbial, Protein Conformation, Time Factors, Technology, Medical and Health Sciences, Developmental Biology, Biological sciences
Abstract

The European XFEL (EuXFEL) is a 3.4-km long X-ray source, which produces femtosecond, ultrabrilliant and spatially coherent X-ray pulses at megahertz (MHz) repetition rates. This X-ray source has been designed to enable the observation of ultrafast processes with near-atomic spatial resolution. Time-resolved crystallographic investigations on biological macromolecules belong to an important class of experiments that explore fundamental and functional structural displacements in these molecules. Due to the unusual MHz X-ray pulse structure at the EuXFEL, these experiments are challenging. Here, we demonstrate how a biological reaction can be followed on ultrafast timescales at the EuXFEL. We investigate the picosecond time range in the photocycle of photoactive yellow protein (PYP) with MHz X-ray pulse rates. We show that difference electron density maps of excellent quality can be obtained. The results connect the previously explored femtosecond PYP dynamics to timescales accessible at synchrotrons. This opens the door to a wide range of time-resolved studies at the EuXFEL.

Published
2020

5. Author Correction: Membrane protein megahertz crystallography at the European XFEL

Author

Gisriel, Chris, Coe, Jesse, Letrun, Romain, Yefanov, Oleksandr M, Luna-Chavez, Cesar, Stander, Natasha E, Lisova, Stella, Mariani, Valerio, Kuhn, Manuela, Aplin, Steve, Grant, Thomas D, Dörner, Katerina, Sato, Tokushi, Echelmeier, Austin, Villarreal, Jorvani Cruz, Hunter, Mark S, Wiedorn, Max O, Knoska, Juraj, Mazalova, Victoria, Roy-Chowdhury, Shatabdi, Yang, Jay-How, Jones, Alex, Bean, Richard, Bielecki, Johan, Kim, Yoonhee, Mills, Grant, Weinhausen, Britta, Meza, Jose D, Al-Qudami, Nasser, Bajt, Saša, Brehm, Gerrit, Botha, Sabine, Boukhelef, Djelloul, Brockhauser, Sandor, Bruce, Barry D, Coleman, Matthew A, Danilevski, Cyril, Discianno, Erin, Dobson, Zachary, Fangohr, Hans, Martin-Garcia, Jose M, Gevorkov, Yaroslav, Hauf, Steffen, Hosseinizadeh, Ahmad, Januschek, Friederike, Ketawala, Gihan K, Kupitz, Christopher, Maia, Luis, Manetti, Maurizio, Messerschmidt, Marc, Michelat, Thomas, Mondal, Jyotirmoy, Ourmazd, Abbas, Previtali, Gianpietro, Sarrou, Iosifina, Schön, Silvan, Schwander, Peter, Shelby, Megan L, Silenzi, Alessandro, Sztuk-Dambietz, Jolanta, Szuba, Janusz, Turcato, Monica, White, Thomas A, Wrona, Krzysztof, Xu, Chen, Abdellatif, Mohamed H, Zook, James D, Spence, John CH, Chapman, Henry N, Barty, Anton, Kirian, Richard A, Frank, Matthias, Ros, Alexandra, Schmidt, Marius, Fromme, Raimund, Mancuso, Adrian P, Fromme, Petra, and Zatsepin, Nadia A

Subjects
Biochemistry and Cell Biology, Biological Sciences
Abstract

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Published
2020

6. Membrane protein megahertz crystallography at the European XFEL

Author

Gisriel, Chris, Coe, Jesse, Letrun, Romain, Yefanov, Oleksandr M, Luna-Chavez, Cesar, Stander, Natasha E, Lisova, Stella, Mariani, Valerio, Kuhn, Manuela, Aplin, Steve, Grant, Thomas D, Dörner, Katerina, Sato, Tokushi, Echelmeier, Austin, Cruz Villarreal, Jorvani, Hunter, Mark S, Wiedorn, Max O, Knoska, Juraj, Mazalova, Victoria, Roy-Chowdhury, Shatabdi, Yang, Jay-How, Jones, Alex, Bean, Richard, Bielecki, Johan, Kim, Yoonhee, Mills, Grant, Weinhausen, Britta, Meza, Jose D, Al-Qudami, Nasser, Bajt, Saša, Brehm, Gerrit, Botha, Sabine, Boukhelef, Djelloul, Brockhauser, Sandor, Bruce, Barry D, Coleman, Matthew A, Danilevski, Cyril, Discianno, Erin, Dobson, Zachary, Fangohr, Hans, Martin-Garcia, Jose M, Gevorkov, Yaroslav, Hauf, Steffen, Hosseinizadeh, Ahmad, Januschek, Friederike, Ketawala, Gihan K, Kupitz, Christopher, Maia, Luis, Manetti, Maurizio, Messerschmidt, Marc, Michelat, Thomas, Mondal, Jyotirmoy, Ourmazd, Abbas, Previtali, Gianpietro, Sarrou, Iosifina, Schön, Silvan, Schwander, Peter, Shelby, Megan L, Silenzi, Alessandro, Sztuk-Dambietz, Jolanta, Szuba, Janusz, Turcato, Monica, White, Thomas A, Wrona, Krzysztof, Xu, Chen, Abdellatif, Mohamed H, Zook, James D, Spence, John CH, Chapman, Henry N, Barty, Anton, Kirian, Richard A, Frank, Matthias, Ros, Alexandra, Schmidt, Marius, Fromme, Raimund, Mancuso, Adrian P, Fromme, Petra, and Zatsepin, Nadia A

Subjects
Biochemistry and Cell Biology, Biological Sciences, 1.1 Normal biological development and functioning, Underpinning research, Generic health relevance, Crystallography, Cyanobacteria, Electrons, Lasers, Membrane Proteins, Models, Molecular, Photosystem I Protein Complex, Static Electricity, Synchrotrons, Thermosynechococcus, X-Rays
Abstract

The world's first superconducting megahertz repetition rate hard X-ray free-electron laser (XFEL), the European XFEL, began operation in 2017, featuring a unique pulse train structure with 886 ns between pulses. With its rapid pulse rate, the European XFEL may alleviate some of the increasing demand for XFEL beamtime, particularly for membrane protein serial femtosecond crystallography (SFX), leveraging orders-of-magnitude faster data collection. Here, we report the first membrane protein megahertz SFX experiment, where we determined a 2.9 Å-resolution SFX structure of the large membrane protein complex, Photosystem I, a > 1 MDa complex containing 36 protein subunits and 381 cofactors. We address challenges to megahertz SFX for membrane protein complexes, including growth of large quantities of crystals and the large molecular and unit cell size that influence data collection and analysis. The results imply that megahertz crystallography could have an important impact on structure determination of large protein complexes with XFELs.

Published
2019

7. Enzyme intermediates captured “on the fly” by mix-and-inject serial crystallography

Author

Olmos, Jose L, Pandey, Suraj, Martin-Garcia, Jose M, Calvey, George, Katz, Andrea, Knoska, Juraj, Kupitz, Christopher, Hunter, Mark S, Liang, Mengning, Oberthuer, Dominik, Yefanov, Oleksandr, Wiedorn, Max, Heyman, Michael, Holl, Mark, Pande, Kanupriya, Barty, Anton, Miller, Mitchell D, Stern, Stephan, Roy-Chowdhury, Shatabdi, Coe, Jesse, Nagaratnam, Nirupa, Zook, James, Verburgt, Jacob, Norwood, Tyler, Poudyal, Ishwor, Xu, David, Koglin, Jason, Seaberg, Matthew H, Zhao, Yun, Bajt, Saša, Grant, Thomas, Mariani, Valerio, Nelson, Garrett, Subramanian, Ganesh, Bae, Euiyoung, Fromme, Raimund, Fung, Russell, Schwander, Peter, Frank, Matthias, White, Thomas A, Weierstall, Uwe, Zatsepin, Nadia, Spence, John, Fromme, Petra, Chapman, Henry N, Pollack, Lois, Tremblay, Lee, Ourmazd, Abbas, Phillips, George N, and Schmidt, Marius

Subjects
Biochemistry and Cell Biology, Biological Sciences, Rare Diseases, Infectious Diseases, Good Health and Well Being, Anti-Bacterial Agents, Bacterial Proteins, Biocatalysis, Ceftriaxone, Cephalosporin Resistance, Crystallography, X-Ray, Kinetics, Lasers, Models, Molecular, Mycobacterium tuberculosis, Time Factors, beta-Lactamases, Developmental Biology
Abstract

BackgroundEver since the first atomic structure of an enzyme was solved, the discovery of the mechanism and dynamics of reactions catalyzed by biomolecules has been the key goal for the understanding of the molecular processes that drive life on earth. Despite a large number of successful methods for trapping reaction intermediates, the direct observation of an ongoing reaction has been possible only in rare and exceptional cases.ResultsHere, we demonstrate a general method for capturing enzyme catalysis "in action" by mix-and-inject serial crystallography (MISC). Specifically, we follow the catalytic reaction of the Mycobacterium tuberculosis β-lactamase with the third-generation antibiotic ceftriaxone by time-resolved serial femtosecond crystallography. The results reveal, in near atomic detail, antibiotic cleavage and inactivation from 30 ms to 2 s.ConclusionsMISC is a versatile and generally applicable method to investigate reactions of biological macromolecules, some of which are of immense biological significance and might be, in addition, important targets for structure-based drug design. With megahertz X-ray pulse rates expected at the Linac Coherent Light Source II and the European X-ray free-electron laser, multiple, finely spaced time delays can be collected rapidly, allowing a comprehensive description of biomolecular reactions in terms of structure and kinetics from the same set of X-ray data.

Published
2018

8. Free-electron laser data for multiple-particle fluctuation scattering analysis.

Author

Pande, Kanupriya, Donatelli, Jeffrey J, Malmerberg, Erik, Foucar, Lutz, Poon, Billy K, Sutter, Markus, Botha, Sabine, Basu, Shibom, Bruce Doak, R, Dörner, Katerina, Epp, Sascha W, Englert, Lars, Fromme, Raimund, Hartmann, Elisabeth, Hartmann, Robert, Hauser, Guenter, Hattne, Johan, Hosseinizadeh, Ahmad, Kassemeyer, Stephan, Lomb, Lukas, Montero, Sebastian F Carron, Menzel, Andreas, Rolles, Daniel, Rudenko, Artem, Seibert, Marvin M, Sierra, Raymond George, Schwander, Peter, Ourmazd, Abbas, Fromme, Petra, Sauter, Nicholas K, Bogan, Michael, Bozek, John, Bostedt, Christoph, Schlichting, Ilme, Kerfeld, Cheryl A, and Zwart, Petrus H

Subjects
Phycodnaviridae, X-Ray Diffraction, Scattering, Small Angle
Abstract

Fluctuation X-ray scattering (FXS) is an emerging experimental technique in which solution scattering data are collected using X-ray exposures below rotational diffusion times, resulting in angularly anisotropic X-ray snapshots that provide several orders of magnitude more information than traditional solution scattering data. Such experiments can be performed using the ultrashort X-ray pulses provided by a free-electron laser source, allowing one to collect a large number of diffraction patterns in a relatively short time. Here, we describe a test data set for FXS, obtained at the Linac Coherent Light Source, consisting of close to 100 000 multi-particle diffraction patterns originating from approximately 50 to 200 Paramecium Bursaria Chlorella virus particles per snapshot. In addition to the raw data, a selection of high-quality pre-processed diffraction patterns and a reference SAXS profile are provided.

Published
2018

9. Minimized Sample Consumption for Time-Resolved Serial Crystallography Applied to the Redox Cycle of Human NQO1

Author

Doppler, Diandra, primary, Grieco, Alice, additional, Koh, Domin, additional, Manna, Abhik, additional, Ansari, Adil, additional, Alvarez, Roberto, additional, Karpos, Konstantinos, additional, Le, Hung, additional, Sonker, Mukul, additional, Ketawala, Gihan, additional, Mahmud, Samira, additional, Quereda Moraleda, Isabel, additional, Pey, Angel Luis, additional, Letrun, Roman, additional, Doerner, Katerina, additional, Koliyadu, Jayanath C. P., additional, de Wijn, Raphael, additional, Bielecki, Johan, additional, Han, Huijong, additional, Kim, Chan, additional, Koua, Faisal, additional, Round, Adam, additional, Sarma, Abhisakh, additional, Sato, Tokushi, additional, Schmidt, Christina, additional, Vakili, Mohammad, additional, Zabelskii, Dmitrii, additional, Bean, Richard, additional, Mancuso, Adrian P., additional, Schulz, Joachim, additional, Fromme, Raimund, additional, Medina, Milagros, additional, Grant, Thomas D, additional, Fromme, Petra, additional, Kirian, Richard A., additional, Botha, Sabine, additional, Martin Garcia, Jose Manuel, additional, and Ros, Alexandra, additional

Published
2024
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10. Structural enzymology using X-ray free electron lasers

Author

Kupitz, Christopher, Olmos, Jose L, Holl, Mark, Tremblay, Lee, Pande, Kanupriya, Pandey, Suraj, Oberthür, Dominik, Hunter, Mark, Liang, Mengning, Aquila, Andrew, Tenboer, Jason, Calvey, George, Katz, Andrea, Chen, Yujie, Wiedorn, Max O, Knoska, Juraj, Meents, Alke, Majriani, Valerio, Norwood, Tyler, Poudyal, Ishwor, Grant, Thomas, Miller, Mitchell D, Xu, Weijun, Tolstikova, Aleksandra, Morgan, Andrew, Metz, Markus, Martin-Garcia, Jose M, Zook, James D, Roy-Chowdhury, Shatabdi, Coe, Jesse, Nagaratnam, Nirupa, Meza, Domingo, Fromme, Raimund, Basu, Shibom, Frank, Matthias, White, Thomas, Barty, Anton, Bajt, Sasa, Yefanov, Oleksandr, Chapman, Henry N, Zatsepin, Nadia, Nelson, Garrett, Weierstall, Uwe, Spence, John, Schwander, Peter, Pollack, Lois, Fromme, Petra, Ourmazd, Abbas, Phillips, George N, and Schmidt, Marius

Subjects
Inorganic Chemistry, Chemical Sciences, Physical Sciences, Infectious Diseases, Good Health and Well Being, Physical chemistry, Condensed matter physics, Particle and high energy physics
Abstract

Mix-and-inject serial crystallography (MISC) is a technique designed to image enzyme catalyzed reactions in which small protein crystals are mixed with a substrate just prior to being probed by an X-ray pulse. This approach offers several advantages over flow cell studies. It provides (i) room temperature structures at near atomic resolution, (ii) time resolution ranging from microseconds to seconds, and (iii) convenient reaction initiation. It outruns radiation damage by using femtosecond X-ray pulses allowing damage and chemistry to be separated. Here, we demonstrate that MISC is feasible at an X-ray free electron laser by studying the reaction of M. tuberculosis ß-lactamase microcrystals with ceftriaxone antibiotic solution. Electron density maps of the apo-ß-lactamase and of the ceftriaxone bound form were obtained at 2.8 Å and 2.4 Å resolution, respectively. These results pave the way to study cyclic and non-cyclic reactions and represent a new field of time-resolved structural dynamics for numerous substrate-triggered biological reactions.

Published
2017

11. Femtosecond structural dynamics drives the trans/cis isomerization in photoactive yellow protein

Author

Pande, Kanupriya, Hutchison, Christopher DM, Groenhof, Gerrit, Aquila, Andy, Robinson, Josef S, Tenboer, Jason, Basu, Shibom, Boutet, Sébastien, DePonte, Daniel P, Liang, Mengning, White, Thomas A, Zatsepin, Nadia A, Yefanov, Oleksandr, Morozov, Dmitry, Oberthuer, Dominik, Gati, Cornelius, Subramanian, Ganesh, James, Daniel, Zhao, Yun, Koralek, Jake, Brayshaw, Jennifer, Kupitz, Christopher, Conrad, Chelsie, Roy-Chowdhury, Shatabdi, Coe, Jesse D, Metz, Markus, Xavier, Paulraj Lourdu, Grant, Thomas D, Koglin, Jason E, Ketawala, Gihan, Fromme, Raimund, Šrajer, Vukica, Henning, Robert, Spence, John CH, Ourmazd, Abbas, Schwander, Peter, Weierstall, Uwe, Frank, Matthias, Fromme, Petra, Barty, Anton, Chapman, Henry N, Moffat, Keith, van Thor, Jasper J, and Schmidt, Marius

Subjects
Biological Sciences, Chemical Sciences, Theoretical and Computational Chemistry, Bacterial Proteins, Crystallography, Isomerism, Light, Photochemical Processes, Photons, Photoreceptors, Microbial, Protein Conformation, Time Factors, General Science & Technology
Abstract

A variety of organisms have evolved mechanisms to detect and respond to light, in which the response is mediated by protein structural changes after photon absorption. The initial step is often the photoisomerization of a conjugated chromophore. Isomerization occurs on ultrafast time scales and is substantially influenced by the chromophore environment. Here we identify structural changes associated with the earliest steps in the trans-to-cis isomerization of the chromophore in photoactive yellow protein. Femtosecond hard x-ray pulses emitted by the Linac Coherent Light Source were used to conduct time-resolved serial femtosecond crystallography on photoactive yellow protein microcrystals over a time range from 100 femtoseconds to 3 picoseconds to determine the structural dynamics of the photoisomerization reaction.

Published
2016

12. Snapshot of an oxygen intermediate in the catalytic reaction of cytochrome c oxidase

Author

Ishigami, Izumi, Lewis-Ballester, Ariel, Echelmeier, Austin, Brehm, Gerrit, Zatsepin, Nadia A., Grant, Thomas D., Coe, Jesse D., Lisova, Stella, Nelson, Garrett, Zhang, Shangji, Dobson, Zachary F., Boutet, Sébastien, Sierra, Raymond G., Batyuk, Alexander, Fromme, Petra, Fromme, Raimund, Spence, John C. H., Ros, Alexandra, Yeh, Syun-Ru, and Rousseau, Denis L.

Published
2019

13. Modular droplet injector for sample conservation providing new structural insight for the conformational heterogeneity in the disease-associated NQO1 enzyme

Author

National Science Foundation (US), National Institutes of Health (US), Department of Energy (US), Max Planck Institute for Medical Research, Comunidad de Madrid, Ministerio de Ciencia, Innovación y Universidades (España), Junta de Andalucía, Doppler, Diandra [0000-0002-3586-3936], Sonker, Mukul [0000-0001-5816-8410], Egatz-Gomez, Ana [0000-0002-6080-943X], Manna, Abhik [0000-0001-8681-6702], Cruz Villarreal, Jorvani [0000-0002-6762-7348], Pey, Angel L. [0000-0001-7706-3243], Ruiz-Fresneda, Miguel A. [0000-0001-6349-6566], Pacheco-Garcia, Juan Luis [0000-0001-6043-7536], Hermoso, Juan A. [0000-0002-1862-8950], Batyuk, Alexander [0000-0002-9393-2880], Fromme, Raimund [0000-0003-4835-1080], Kirian, Richard A. [0000-0001-7197-3086], Martín-García, José M. [0000-0002-4558-3858], Ros, Alexandra [0000-0001-7709-8331], Doppler, Diandra, Sonker, Mukul, Egatz-Gomez, Ana, Grieco, Alice, Zaare, Sahba, Jernigan, Rebecca, Meza-Aguilar, Jose Domingo, Rabbani, Mohammad T., Manna, Abhik, Alvarez, Roberto C., Karpos, Konstantinos, Cruz Villarreal, Jorvani, Nelson, Garrett, Yang, Jay-How, Carrion, Jackson, Morin, Katherine, Ketawala, Gihan K, Pey, Angel L., Ruiz-Fresneda, Miguel A., Pacheco-Garcia, Juan Luis, Hermoso, Juan A., Nazari, Reza, Sierra, Raymond, Hunter, Mark S., Batyuk, Alexander, Kupitz, Christopher J., Sublett, Robert E., Lisova, Stella, Mariani, Valerio, Boutet, Sébastien, Fromme, Raimund, Grant, Thomas D., Botha, Sabine, Fromme, Petra, Kirian, Richard A., Martín-García, José M., Ros, Alexandra, National Science Foundation (US), National Institutes of Health (US), Department of Energy (US), Max Planck Institute for Medical Research, Comunidad de Madrid, Ministerio de Ciencia, Innovación y Universidades (España), Junta de Andalucía, Doppler, Diandra [0000-0002-3586-3936], Sonker, Mukul [0000-0001-5816-8410], Egatz-Gomez, Ana [0000-0002-6080-943X], Manna, Abhik [0000-0001-8681-6702], Cruz Villarreal, Jorvani [0000-0002-6762-7348], Pey, Angel L. [0000-0001-7706-3243], Ruiz-Fresneda, Miguel A. [0000-0001-6349-6566], Pacheco-Garcia, Juan Luis [0000-0001-6043-7536], Hermoso, Juan A. [0000-0002-1862-8950], Batyuk, Alexander [0000-0002-9393-2880], Fromme, Raimund [0000-0003-4835-1080], Kirian, Richard A. [0000-0001-7197-3086], Martín-García, José M. [0000-0002-4558-3858], Ros, Alexandra [0000-0001-7709-8331], Doppler, Diandra, Sonker, Mukul, Egatz-Gomez, Ana, Grieco, Alice, Zaare, Sahba, Jernigan, Rebecca, Meza-Aguilar, Jose Domingo, Rabbani, Mohammad T., Manna, Abhik, Alvarez, Roberto C., Karpos, Konstantinos, Cruz Villarreal, Jorvani, Nelson, Garrett, Yang, Jay-How, Carrion, Jackson, Morin, Katherine, Ketawala, Gihan K, Pey, Angel L., Ruiz-Fresneda, Miguel A., Pacheco-Garcia, Juan Luis, Hermoso, Juan A., Nazari, Reza, Sierra, Raymond, Hunter, Mark S., Batyuk, Alexander, Kupitz, Christopher J., Sublett, Robert E., Lisova, Stella, Mariani, Valerio, Boutet, Sébastien, Fromme, Raimund, Grant, Thomas D., Botha, Sabine, Fromme, Petra, Kirian, Richard A., Martín-García, José M., and Ros, Alexandra

Abstract

Droplet injection strategies are a promising tool to reduce the large amount of sample consumed in serial femtosecond crystallography (SFX) measurements at X-ray free electron lasers (XFELs) with continuous injection approaches. Here, we demonstrate a new modular microfluidic droplet injector (MDI) design that was successfully applied to deliver microcrystals of the human NAD(P)H:quinone oxidoreductase 1 (NQO1) and phycocyanin. We investigated droplet generation conditions through electrical stimulation for both protein samples and implemented hardware and software components for optimized crystal injection at the Macromolecular Femtosecond Crystallography (MFX) instrument at the Stanford Linac Coherent Light Source (LCLS). Under optimized droplet injection conditions, we demonstrate that up to 4-fold sample consumption savings can be achieved with the droplet injector. In addition, we collected a full data set with droplet injection for NQO1 protein crystals with a resolution up to 2.7 Å, leading to the first room-temperature structure of NQO1 at an XFEL. NQO1 is a flavoenzyme associated with cancer, Alzheimer's and Parkinson's disease, making it an attractive target for drug discovery. Our results reveal for the first time that residues Tyr128 and Phe232, which play key roles in the function of the protein, show an unexpected conformational heterogeneity at room temperature within the crystals. These results suggest that different substates exist in the conformational ensemble of NQO1 with functional and mechanistic implications for the enzyme's negative cooperativity through a conformational selection mechanism. Our study thus demonstrates that microfluidic droplet injection constitutes a robust sample-conserving injection method for SFX studies on protein crystals that are difficult to obtain in amounts necessary for continuous injection, including the large sample quantities required for time-resolved mix-and-inject studies.

Published
2023

14. Crystal structure of rhodopsin bound to arrestin by femtosecond X-ray laser

Author

Kang, Yanyong, Zhou, X Edward, Gao, Xiang, He, Yuanzheng, Liu, Wei, Ishchenko, Andrii, Barty, Anton, White, Thomas A, Yefanov, Oleksandr, Han, Gye Won, Xu, Qingping, de Waal, Parker W, Ke, Jiyuan, Tan, MH Eileen, Zhang, Chenghai, Moeller, Arne, West, Graham M, Pascal, Bruce D, Van Eps, Ned, Caro, Lydia N, Vishnivetskiy, Sergey A, Lee, Regina J, Suino-Powell, Kelly M, Gu, Xin, Pal, Kuntal, Ma, Jinming, Zhi, Xiaoyong, Boutet, Sébastien, Williams, Garth J, Messerschmidt, Marc, Gati, Cornelius, Zatsepin, Nadia A, Wang, Dingjie, James, Daniel, Basu, Shibom, Roy-Chowdhury, Shatabdi, Conrad, Chelsie E, Coe, Jesse, Liu, Haiguang, Lisova, Stella, Kupitz, Christopher, Grotjohann, Ingo, Fromme, Raimund, Jiang, Yi, Tan, Minjia, Yang, Huaiyu, Li, Jun, Wang, Meitian, Zheng, Zhong, Li, Dianfan, Howe, Nicole, Zhao, Yingming, Standfuss, Jörg, Diederichs, Kay, Dong, Yuhui, Potter, Clinton S, Carragher, Bridget, Caffrey, Martin, Jiang, Hualiang, Chapman, Henry N, Spence, John CH, Fromme, Petra, Weierstall, Uwe, Ernst, Oliver P, Katritch, Vsevolod, Gurevich, Vsevolod V, Griffin, Patrick R, Hubbell, Wayne L, Stevens, Raymond C, Cherezov, Vadim, Melcher, Karsten, and Xu, H Eric

Subjects
1.1 Normal biological development and functioning, Underpinning research, Generic health relevance, Animals, Arrestin, Binding Sites, Crystallography, X-Ray, Disulfides, Humans, Lasers, Mice, Models, Molecular, Multiprotein Complexes, Protein Binding, Reproducibility of Results, Rhodopsin, Signal Transduction, X-Rays, General Science & Technology
Abstract

G-protein-coupled receptors (GPCRs) signal primarily through G proteins or arrestins. Arrestin binding to GPCRs blocks G protein interaction and redirects signalling to numerous G-protein-independent pathways. Here we report the crystal structure of a constitutively active form of human rhodopsin bound to a pre-activated form of the mouse visual arrestin, determined by serial femtosecond X-ray laser crystallography. Together with extensive biochemical and mutagenesis data, the structure reveals an overall architecture of the rhodopsin-arrestin assembly in which rhodopsin uses distinct structural elements, including transmembrane helix 7 and helix 8, to recruit arrestin. Correspondingly, arrestin adopts the pre-activated conformation, with a ∼20° rotation between the amino and carboxy domains, which opens up a cleft in arrestin to accommodate a short helix formed by the second intracellular loop of rhodopsin. This structure provides a basis for understanding GPCR-mediated arrestin-biased signalling and demonstrates the power of X-ray lasers for advancing the frontiers of structural biology.

Published
2015

15. Time-resolved protein nanocrystallography using an X-ray free-electron laser

Author

Aquila, Andrew, Hunter, Mark S, Doak, R Bruce, Kirian, Richard A, Fromme, Petra, White, Thomas A, Andreasson, Jakob, Arnlund, David, Bajt, Saša, Barends, Thomas RM, Barthelmess, Miriam, Bogan, Michael J, Bostedt, Christoph, Bottin, Hervé, Bozek, John D, Caleman, Carl, Coppola, Nicola, Davidsson, Jan, DePonte, Daniel P, Elser, Veit, Epp, Sascha W, Erk, Benjamin, Fleckenstein, Holger, Foucar, Lutz, Frank, Matthias, Fromme, Raimund, Graafsma, Heinz, Grotjohann, Ingo, Gumprecht, Lars, Hajdu, Janos, Hampton, Christina Y, Hartmann, Andreas, Hartmann, Robert, Hau-Riege, Stefan, Hauser, Günter, Hirsemann, Helmut, Holl, Peter, Holton, James M, Hömke, André, Johansson, Linda, Kimmel, Nils, Kassemeyer, Stephan, Krasniqi, Faton, Kühnel, Kai-Uwe, Liang, Mengning, Lomb, Lukas, Malmerberg, Erik, Marchesini, Stefano, Martin, Andrew V, Maia, Filipe RNC, Messerschmidt, Marc, Nass, Karol, Reich, Christian, Neutze, Richard, Rolles, Daniel, Rudek, Benedikt, Rudenko, Artem, Schlichting, Ilme, Schmidt, Carlo, Schmidt, Kevin E, Schulz, Joachim, Seibert, M Marvin, Shoeman, Robert L, Sierra, Raymond, Soltau, Heike, Starodub, Dmitri, Stellato, Francesco, Stern, Stephan, Strüder, Lothar, Timneanu, Nicusor, Ullrich, Joachim, Wang, Xiaoyu, Williams, Garth J, Weidenspointner, Georg, Weierstall, Uwe, Wunderer, Cornelia, Barty, Anton, Spence, John CH, and Chapman, Henry N

Subjects
Crystallography, X-Ray, Electrons, Ferredoxins, Lasers, Nanostructures, Protein Conformation, X-Ray Diffraction, X-Rays, Optical Physics, Electrical and Electronic Engineering, Communications Technologies, Optics
Abstract

We demonstrate the use of an X-ray free electron laser synchronized with an optical pump laser to obtain X-ray diffraction snapshots from the photoactivated states of large membrane protein complexes in the form of nanocrystals flowing in a liquid jet. Light-induced changes of Photosystem I-Ferredoxin co-crystals were observed at time delays of 5 to 10 µs after excitation. The result correlates with the microsecond kinetics of electron transfer from Photosystem I to ferredoxin. The undocking process that follows the electron transfer leads to large rearrangements in the crystals that will terminally lead to the disintegration of the crystals. We describe the experimental setup and obtain the first time-resolved femtosecond serial X-ray crystallography results from an irreversible photo-chemical reaction at the Linac Coherent Light Source. This technique opens the door to time-resolved structural studies of reaction dynamics in biological systems.

Published
2012

16. Segmented flow generator for serial crystallography at the European X-ray free electron laser

Author

Echelmeier, Austin, Cruz Villarreal, Jorvani, Messerschmidt, Marc, Kim, Daihyun, Coe, Jesse D., Thifault, Darren, Botha, Sabine, Egatz-Gomez, Ana, Gandhi, Sahir, Brehm, Gerrit, Conrad, Chelsie E., Hansen, Debra T., Madsen, Caleb, Bajt, Saša, Meza-Aguilar, J. Domingo, Oberthür, Dominik, Wiedorn, Max O., Fleckenstein, Holger, Mendez, Derek, Knoška, Juraj, Martin-Garcia, Jose M., Hu, Hao, Lisova, Stella, Allahgholi, Aschkan, Gevorkov, Yaroslav, Ayyer, Kartik, Aplin, Steve, Ginn, Helen Mary, Graafsma, Heinz, Morgan, Andrew J., Greiffenberg, Dominic, Klujev, Alexander, Laurus, Torsten, Poehlsen, Jennifer, Trunk, Ulrich, Mezza, Davide, Schmidt, Bernd, Kuhn, Manuela, Fromme, Raimund, Sztuk-Dambietz, Jolanta, Raab, Natascha, Hauf, Steffen, Silenzi, Alessandro, Michelat, Thomas, Xu, Chen, Danilevski, Cyril, Parenti, Andrea, Mekinda, Leonce, Weinhausen, Britta, Mills, Grant, Vagovic, Patrik, Kim, Yoonhee, Kirkwood, Henry, Bean, Richard, Bielecki, Johan, Stern, Stephan, Giewekemeyer, Klaus, Round, Adam R., Schulz, Joachim, Dörner, Katerina, Grant, Thomas D., Mariani, Valerio, Barty, Anton, Mancuso, Adrian P., Weierstall, Uwe, Spence, John C. H., Chapman, Henry N., Zatsepin, Nadia, Fromme, Petra, Kirian, Richard A., and Ros, Alexandra

Published
2020
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19. Atomic structure of granulin determined from native nanocrystalline granulovirus using an X-ray free-electron laser

Author

Gati, Cornelius, Oberthuer, Dominik, Yefanov, Oleksandr, Bunker, Richard D., Stellato, Francesco, Chiu, Elaine, Yeh, Shin-Mei, Aquila, Andrew, Basu, Shibom, Bean, Richard, Beyerlein, Kenneth R., Botha, Sabine, Boutet, Sébastien, DePonte, Daniel P., Doak, R. Bruce, Fromme, Raimund, Galli, Lorenzo, Grotjohann, Ingo, James, Daniel R., Kupitz, Christopher, Lomb, Lukas, Messerschmidt, Marc, Nass, Karol, Rendek, Kimberly, Shoeman, Robert L., Wang, Dingjie, Weierstall, Uwe, White, Thomas A., Williams, Garth J., Zatsepin, Nadia A., Fromme, Petra, Spence, John C. H., Goldie, Kenneth N., Jehle, Johannes A., Metcalf, Peter, Barty, Anton, and Chapman, Henry N.

Published
2017

22. Detection of a geminate photoproduct of bovine cytochrome c oxidase by time-resolved serial femtosecond crystallography

Author

Ishigami, Izumi, primary, Carbajo, Sergio, additional, Zatsepin, Nadia, additional, Hikita, Masahide, additional, Conrad, Chelsie E., additional, Nelson, Garrett, additional, Coe, Jesse, additional, Basu, Shibom, additional, Grant, Thomas, additional, Seaberg, Matthew H., additional, Sierra, Raymond G., additional, Hunter, Mark S., additional, Fromme, Petra, additional, Fromme, Raimund, additional, Rousseau, Denis L., additional, and Yeh, Syun-Ru, additional

Published
2023
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24. Modular droplet injector for sample conservation providing new structural insight for the conformational heterogeneity in the disease-associated NQO1 enzyme

Author

Doppler, Diandra, primary, Sonker, Mukul, additional, Egatz-Gomez, Ana, additional, Grieco, Alice, additional, Zaare, Sahba, additional, Jernigan, Rebecca, additional, Meza-Aguilar, Jose Domingo, additional, Rabbani, Mohammad T., additional, Manna, Abhik, additional, Alvarez, Roberto C., additional, Karpos, Konstantinos, additional, Cruz Villarreal, Jorvani, additional, Nelson, Garrett, additional, Yang, Jay-How, additional, Carrion, Jackson, additional, Morin, Katherine, additional, Ketawala, Gihan K., additional, Pey, Angel L., additional, Ruiz-Fresneda, Miguel Angel, additional, Pacheco-Garcia, Juan Luis, additional, Hermoso, Juan A., additional, Nazari, Reza, additional, Sierra, Raymond, additional, Hunter, Mark S., additional, Batyuk, Alexander, additional, Kupitz, Christopher J., additional, Sublett, Robert E., additional, Lisova, Stella, additional, Mariani, Valerio, additional, Boutet, Sébastien, additional, Fromme, Raimund, additional, Grant, Thomas D., additional, Botha, Sabine, additional, Fromme, Petra, additional, Kirian, Richard A., additional, Martin-Garcia, Jose Manuel, additional, and Ros, Alexandra, additional

Published
2023
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25. Physachenolide C is a Potent, Selective BET Inhibitor

Author

Zerio, Christopher J., primary, Sivinski, Jared, additional, Wijeratne, E. M. Kithsiri, additional, Xu, Ya-Ming, additional, Ngo, Duc T., additional, Ambrose, Andrew J., additional, Villa-Celis, Luis, additional, Ghadirian, Niloofar, additional, Clarkson, Michael W., additional, Zhang, Donna D., additional, Horton, Nancy C., additional, Gunatilaka, A. A. Leslie, additional, Fromme, Raimund, additional, and Chapman, Eli, additional

Published
2022
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27. Electrically stimulated droplet injector for reduced sample consumption in serial crystallography

Author

Sonker, Mukul, primary, Doppler, Diandra, additional, Egatz-Gomez, Ana, additional, Zaare, Sahba, additional, Rabbani, Mohammad T., additional, Manna, Abhik, additional, Cruz Villarreal, Jorvani, additional, Nelson, Garrett, additional, Ketawala, Gihan K., additional, Karpos, Konstantinos, additional, Alvarez, Roberto C., additional, Nazari, Reza, additional, Thifault, Darren, additional, Jernigan, Rebecca, additional, Oberthür, Dominik, additional, Han, Huijong, additional, Sierra, Raymond, additional, Hunter, Mark S., additional, Batyuk, Alexander, additional, Kupitz, Christopher J., additional, Sublett, Robert E., additional, Poitevin, Frederic, additional, Lisova, Stella, additional, Mariani, Valerio, additional, Tolstikova, Alexandra, additional, Boutet, Sebastien, additional, Messerschmidt, Marc, additional, Meza-Aguilar, J. Domingo, additional, Fromme, Raimund, additional, Martin-Garcia, Jose M., additional, Botha, Sabine, additional, Fromme, Petra, additional, Grant, Thomas D., additional, Kirian, Richard A., additional, and Ros, Alexandra, additional

Published
2022
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28. Time-resolved serial crystallography captures high-resolution intermediates of photoactive yellow protein

Author

Tenboer, Jason, Basu, Shibom, Zatsepin, Nadia, Pande, Kanupriya, Milathianaki, Despina, Frank, Matthias, Hunter, Mark, Boutet, Sébastien, Williams, Garth J., Koglin, Jason E., Oberthuer, Dominik, Heymann, Michael, Kupitz, Christopher, Conrad, Chelsie, Coe, Jesse, Roy-Chowdhury, Shatabdi, Weierstall, Uwe, James, Daniel, Wang, Dingjie, Grant, Thomas, Barty, Anton, Yefanov, Oleksandr, Scales, Jennifer, Gati, Cornelius, Seuring, Carolin, Srajer, Vukica, Henning, Robert, Schwander, Peter, Fromme, Raimund, Ourmazd, Abbas, Moffat, Keith, Van Thor, Jasper J., Spence, John C. H., Fromme, Petra, Chapman, Henry N., and Schmidt, Marius

Published
2014

30. Structural and biophysical properties of FopA, a major outer membrane protein of Francisella tularensis

Author

Nagaratnam, Nirupa, primary, Martin-Garcia, Jose M., additional, Yang, Jay-How, additional, Goode, Matthew R., additional, Ketawala, Gihan, additional, Craciunescu, Felicia M., additional, Zook, James D., additional, Sonowal, Manashi, additional, Williams, Dewight, additional, Grant, Thomas D., additional, Fromme, Raimund, additional, Hansen, Debra T., additional, and Fromme, Petra, additional

Published
2022
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32. Serial Femtosecond Crystallography of G Protein-Coupled Receptors

Author

Liu, Wei, Wacker, Daniel, Gati, Cornelius, Han, Gye Won, James, Daniel, Wang, Dingjie, Nelson, Garrett, Weierstall, Uwe, Katritch, Vsevolod, Barty, Anton, Zatsepin, Nadia A., Li, Dianfan, Messerschmidt, Marc, Boutet, Sébastien, Williams, Garth J., Koglin, Jason E., Seibert, M. Marvin, Wang, Chong, Shah, Syed T. A., Basu, Shibom, Fromme, Raimund, Kupitz, Christopher, Rendek, Kimberley N., Grotjohann, Ingo, Fromme, Petra, Kirian, Richard A., Beyerlein, Kenneth R., White, Thomas A., Chapman, Henry N., Caffrey, Martin, Spence, John C. H., Stevens, Raymond C., and Cherezov, Vadim

Published
2013
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33. Detection of a Geminate Photoproduct of Bovine Cytochrome cOxidase by Time-Resolved Serial Femtosecond Crystallography

Author

Ishigami, Izumi, Carbajo, Sergio, Zatsepin, Nadia, Hikita, Masahide, Conrad, Chelsie E., Nelson, Garrett, Coe, Jesse, Basu, Shibom, Grant, Thomas, Seaberg, Matthew H., Sierra, Raymond G., Hunter, Mark S., Fromme, Petra, Fromme, Raimund, Rousseau, Denis L., and Yeh, Syun-Ru

Abstract

Cytochrome coxidase (CcO) is a large membrane-bound hemeprotein that catalyzes the reduction of dioxygen to water. Unlike classical dioxygen binding hemeproteins with a heme bgroup in their active sites, CcO has a unique binuclear center (BNC) composed of a copper atom (CuB) and a heme a3iron, where O2binds and is reduced to water. CO is a versatile O2surrogate in ligand binding and escape reactions. Previous time-resolved spectroscopic studies of the CO complexes of bovine CcO (bCcO) revealed that photolyzing CO from the heme a3iron leads to a metastable intermediate (CuB-CO), where CO is bound to CuB, before it escapes out of the BNC. Here, with a pump-probe based time-resolved serial femtosecond X-ray crystallography, we detected a geminate photoproduct of the bCcO–CO complex, where CO is dissociated from the heme a3iron and moved to a temporary binding site midway between the CuBand the heme a3iron, while the locations of the two metal centers and the conformation of Helix-X, housing the proximal histidine ligand of the heme a3iron, remain in the CO complex state. This new structure, combined with other reported structures of bCcO, allows for a clearer definition of the ligand dissociation trajectory as well as the associated protein dynamics.

Published
2023
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34. Natively Inhibited Trypanosoma brucei Cathepsin B Structure Determined by Using an X-ray Laser

Author

Redecke, Lars, Nass, Karol, Deponte, Daniel P., White, Thomas A., Rehders, Dirk, Barty, Anton, Stellato, Francesco, Liang, Mengning, Barends, Thomas R.M., Boutet, Sébastien, Williams, Garth J., Messerschmidt, Marc, Seibert, M. Marvin, Aquila, Andrew, Arnlund, David, Bajt, Sasa, Barth, Torsten, Bogan, Michael J., Caleman, Carl, Chao, Tzu-Chiao, Doak, R. Bruce, Fleckenstein, Holger, Frank, Matthias, Fromme, Raimund, Galli, Lorenzo, Grotjohann, Ingo, Hunter, Mark S., Johansson, Linda C., Kassemeyer, Stephan, Katona, Gergely, Kirian, Richard A., Koopmann, Rudolf, Kupitz, Chris, Lomb, Lukas, Martin, Andrew V., Mogk, Stefan, Neutze, Richard, Shoeman, Robert L., Steinbrener, Jan, Timneanu, Nicusor, Wang, Dingjie, Weierstall, Uwe, Zatsepin, Nadia A., Spence, John C. H., Fromme, Petra, Schlichting, Ilme, Duszenko, Michael, Betzel, Christian, and Chapman, Henry N.

Published
2013
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35. Electrically stimulated droplet injector for reduced sample consumption in serial crystallography

Author

National Science Foundation (US), National Institutes of Health (US), Department of Energy (US), Max Planck Institute for Medical Research, Sonker, Mukul, Doppler, Diandra, Egatz-Gomez, Ana, Zaare, Sahba, Rabbani, Mohammad T., Manna, Abhik, Cruz Villarreal, Jorvani, Nelson, Garrett, Ketawala, Gihan K., Karpos, Konstantinos, Alvarez, Roberto C., Nazari, Reza, Thifault, Darren, Jernigan, Rebecca, Oberthür, Dominik, Han, Huijong, Sierra, Raymond, Hunter, Mark S., Batyuk, Alexander, Kupitz, Christopher J., Sublett, Robert E., Poitevin, Frederic, Lisova, Stella, Mariani, Valerio, Tolstikova, Alexandra, Boutet, Sebastien, Messerschmidt, Marc, Meza-Aguilar, J Domingo, Fromme, Raimund, Martín-García, Jose Manuel, Botha, Sabine, Fromme, Petra, Grant, Thomas D., Kirian, Richard A., Ros, Alexandra, National Science Foundation (US), National Institutes of Health (US), Department of Energy (US), Max Planck Institute for Medical Research, Sonker, Mukul, Doppler, Diandra, Egatz-Gomez, Ana, Zaare, Sahba, Rabbani, Mohammad T., Manna, Abhik, Cruz Villarreal, Jorvani, Nelson, Garrett, Ketawala, Gihan K., Karpos, Konstantinos, Alvarez, Roberto C., Nazari, Reza, Thifault, Darren, Jernigan, Rebecca, Oberthür, Dominik, Han, Huijong, Sierra, Raymond, Hunter, Mark S., Batyuk, Alexander, Kupitz, Christopher J., Sublett, Robert E., Poitevin, Frederic, Lisova, Stella, Mariani, Valerio, Tolstikova, Alexandra, Boutet, Sebastien, Messerschmidt, Marc, Meza-Aguilar, J Domingo, Fromme, Raimund, Martín-García, Jose Manuel, Botha, Sabine, Fromme, Petra, Grant, Thomas D., Kirian, Richard A., and Ros, Alexandra

Abstract

With advances in X-ray free-electron lasers (XFELs), serial femtosecond crystallography (SFX) has enabled the static and dynamic structure determination for challenging proteins such as membrane protein complexes. In SFX with XFELs, the crystals are typically destroyed after interacting with a single XFEL pulse. Therefore, thousands of new crystals must be sequentially introduced into the X-ray beam to collect full data sets. Because of the serial nature of any SFX experiment, up to 99% of the sample delivered to the X-ray beam during its "off-time" between X-ray pulses is wasted due to the intrinsic pulsed nature of all current XFELs. To solve this major problem of large and often limiting sample consumption, we report on improvements of a revolutionary sample-saving method that is compatible with all current XFELs. We previously reported 3D-printed injection devices coupled with gas dynamic virtual nozzles (GDVNs) capable of generating samples containing droplets segmented by an immiscible oil phase for jetting crystal-laden droplets into the path of an XFEL. Here, we have further improved the device design by including metal electrodes inducing electrowetting effects for improved control over droplet generation frequency to stimulate the droplet release to matching the XFEL repetition rate by employing an electrical feedback mechanism. We report the improvements in this electrically triggered segmented flow approach for sample conservation in comparison with a continuous GDVN injection using the microcrystals of lysozyme and 3-deoxy-D-manno-octulosonate 8-phosphate synthase and report the segmented flow approach for sample injection applied at the Macromolecular Femtosecond Crystallography instrument at the Linear Coherent Light Source for the first time.

Published
2022

36. High-Resolution Protein Structure Determination by Serial Femtosecond Crystallography

Author

Boutet, Sébastien, Lomb, Lukas, Williams, Garth J., Barends, Thomas R. M., Aquila, Andrew, Doak, R. Bruce, Weierstall, Uwe, DePonte, Daniel P., Steinbrener, Jan, Shoeman, Robert L., Messerschmidt, Marc, Barty, Anton, White, Thomas A., Kassemeyer, Stephan, Kirian, Richard A., Seibert, M. Marvin, Montanez, Paul A., Kenney, Chris, Herbst, Ryan, Hart, Philip, Pines, Jack, Haller, Gunther, Gruner, Sol M., Philipp, Hugh T., Tate, Mark W., Hromalik, Marianne, Koerner, Lucas J., van Bakel, Niels, Morse, John, Ghonsalves, Wilfred, Arnlund, David, Bogan, Michael J., Caleman, Carl, Fromme, Raimund, Hampton, Christina Y., Hunter, Mark S., Johansson, Linda C., Katona, Gergely, Kupitz, Christopher, Liang, Mengning, Martin, Andrew V., Nass, Karol, Redecke, Lars, Stellato, Francesco, Timneanu, Nicusor, Wang, Dingjie, Zatsepin, Nadia A., Schafer, Donald, Defever, James, Neutze, Richard, Fromme, Petra, Spence, John C. H., Chapman, Henry N., and Schlichting, Ilme

Published
2012
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37. Serial time-resolved crystallography of photosystem II using a femtosecond X-ray laser

Author

Kupitz, Christopher, Basu, Shibom, Grotjohann, Ingo, Fromme, Raimund, Zatsepin, Nadia A., Rendek, Kimberly N., and Hunter, Mark S.

Subjects
Crystallography -- Research, Free electron lasers -- Research, Photosystem II -- Research, Environmental issues, Science and technology, Zoology and wildlife conservation
Abstract

Femtosecond X-ray pulses were used to obtain diffraction data on photosystem II, revealing conformational changes as the complex transitions from the dark S.sub.1 state to the double-pumped S.sub.3 state; the time-resolved serial femtosecond crystallography technique enables structural determination of protein conformations that are highly prone to traditional radiation damage. An X-ray snapshot of photosystem II structure It has recently been shown that extremely short and intense radiation pulses from X-ray free-electron lasers can be used to obtain diffraction data on nanometre- to micrometre-sized protein crystals before the crystal suffers radiation damage. The hope is that this 'serial femtosecond crystallography' (SFX) approach will produce structures of proteins and protein complexes that do not yield well-ordered macroscopic crystals. These authors collected time-resolved SFX data on small crystals of photosystem II of photosynthesis during its transition from the 'dark' S1 state to the double-excited S3 state. At present the resolution of this technique is moderate, but it is sufficient to reveal significant conformational changes at the Mn.sub.4CaO.sub.5 cluster at the heart of the oxygen evolving complex and at the electron acceptor site. Photosynthesis, a process catalysed by plants, algae and cyanobacteria converts sunlight to energy thus sustaining all higher life on Earth. Two large membrane protein complexes, photosystem I and II (PSI and PSII), act in series to catalyse the light-driven reactions in photosynthesis. PSII catalyses the light-driven water splitting process, which maintains the Earth's oxygenic atmosphere.sup.1. In this process, the oxygen-evolving complex (OEC) of PSII cycles through five states, S.sub.0 to S.sub.4, in which four electrons are sequentially extracted from the OEC in four light-driven charge-separation events. Here we describe time resolved experiments on PSII nano/microcrystals from Thermosynechococcus elongatus performed with the recently developed.sup.2 technique of serial femtosecond crystallography. Structures have been determined from PSII in the dark S.sub.1 state and after double laser excitation (putative S.sub.3 state) at 5 and 5.5 Å resolution, respectively. The results provide evidence that PSII undergoes significant conformational changes at the electron acceptor side and at the Mn.sub.4CaO.sub.5 core of the OEC. These include an elongation of the metal cluster, accompanied by changes in the protein environment, which could allow for binding of the second substrate water molecule between the more distant protruding Mn (referred to as the 'dangler' Mn) and the Mn.sub.3CaO.sub.x cubane in the S.sub.2 to S.sub.3 transition, as predicted by spectroscopic and computational studies.sup.3,4. This work shows the great potential for time-resolved serial femtosecond crystallography for investigation of catalytic processes in biomolecules., Author(s): Christopher Kupitz [sup.1] , Shibom Basu [sup.1] , Ingo Grotjohann [sup.1] , Raimund Fromme [sup.1] , Nadia A. Zatsepin [sup.2] , Kimberly N. Rendek [sup.1] , Mark S. Hunter [...]

Published
2014
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38. PROTEIN STRUCTURE: Femtosecond structural dynamics drives the trans/cis isomerization in photoactive yellow protein

Author

Pande, Kanupriya, Hutchison, Christopher D. M., Groenhof, Gerrit, Aquila, Andy, Robinson, Josef S., Tenboer, Jason, Basu, Shibom, Boutet, Sébastien, DePonte, Daniel P., Liang, Mengning, White, Thomas A., Zatsepin, Nadia A., Yefanov, Oleksandr, Morozov, Dmitry, Oberthuer, Dominik, Gati, Cornelius, Subramanian, Ganesh, James, Daniel, Zhao, Yun, Koralek, Jake, Brayshaw, Jennifer, Kupitz, Christopher, Conrad, Chelsie, Roy-Chowdhury, Shatabdi, Coe, Jesse D., Metz, Markus, Xavier, Paulraj Lourdu, Grant, Thomas D., Koglin, Jason E., Ketawala, Gihan, Fromme, Raimund, Šrajer, Vukica, Henning, Robert, Spence, John C. H., Ourmazd, Abbas, Schwander, Peter, Weierstall, Uwe, Frank, Matthias, Fromme, Petra, Barty, Anton, Chapman, Henry N., Moffat, Keith, van Thor, Jasper J., and Schmidt, Marius

Published
2016
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39. Macromolecular diffractive imaging using imperfect crystals

Author

Ayyer, Kartik, Yefanov, Oleksandr M., Oberthür, Dominik, Roy-Chowdhury, Shatabdi, Galli, Lorenzo, Mariani, Valerio, Basu, Shibom, Coe, Jesse, Conrad, Chelsie E., Fromme, Raimund, Schaffer, Alexander, Dörner, Katerina, James, Daniel, Kupitz, Christopher, Metz, Markus, Nelson, Garrett, Xavier, Paulraj Lourdu, Beyerlein, Kenneth R., Schmidt, Marius, Sarrou, Iosifina, Spence, John C. H., Weierstall, Uwe, White, Thomas A., Yang, Jay-How, Zhao, Yun, Liang, Mengning, Aquila, Andrew, Hunter, Mark S., Robinson, Joseph S., Koglin, Jason E., Boutet, Sébastien, Fromme, Petra, Barty, Anton, and Chapman, Henry N.

Published
2016
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40. Physachenolide C is a Potent, Selective BET Inhibitor.

Author

Zerio, Christopher J., Sivinski, Jared, Wijeratne, E. M. Kithsiri, Xu, Ya-Ming, Ngo, Duc T., Ambrose, Andrew J., Villa-Celis, Luis, Ghadirian, Niloofar, Clarkson, Michael W., Zhang, Donna D., Horton, Nancy C., Gunatilaka, A. A. Leslie, Fromme, Raimund, and Chapman, Eli

Published
2023
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42. STRUCTURAL BIOLOGY: Time-resolved serial crystallography captures high-resolution intermediates of photoactive yellow protein

Author

Tenboer, Jason, Basu, Shibom, Zatsepin, Nadia, Pande, Kanupriya, Milathianaki, Despina, Frank, Matthias, Hunter, Mark, Boutet, Sébastien, Williams, Garth J., Koglin, Jason E., Oberthuer, Dominik, Heymann, Michael, Kupitz, Christopher, Conrad, Chelsie, Coe, Jesse, Roy-Chowdhury, Shatabdi, Weierstall, Uwe, James, Daniel, Wang, Dingjie, Grant, Thomas, Barty, Anton, Yefanov, Oleksandr, Scales, Jennifer, Gati, Cornelius, Seuring, Carolin, Srajer, Vukica, Henning, Robert, Schwander, Peter, Fromme, Raimund, Ourmazd, Abbas, Moffat, Keith, Van Thor, Jasper J., Spence, John C. H., Fromme, Petra, Chapman, Henry N., and Schmidt, Marius

Published
2014
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43. Direct Observation of the Mechanism of Antibiotic Resistance by Mix-and-Inject at the European XFEL

Author

Pandey, Suraj, Bean, Richard, Hunter, Mark, Abdellatif, Mohamed H., Bajt, Saša, Bondar, Valerii, Echelmeier, Austin, Doppler, Diandra, Emons, Moritz, Frank, Matthias, Fromme, Raimund, Gevorkov, Yaroslav, Sato, Tokushi, Giovanetti, Gabriele, Jiang, Man, Kim, Daihyun, Kim, Yoonhee, Kirkwood, Henry, Klimovskaia, Anna, Knoska, Juraj, Koua, Faisal H. M., Letrun, Romain, Lisova, Stella, Poudyal, Ishwor, Maia, Luis, Mazalova, Victoria, Meza, Domingo, Michelat, Thomas, Ourmazd, Abbas, Palmer, Guido, Ramilli, Marco, Schubert, Robin, Schwander, Peter, Silenzi, Alessandro, Bielecki, Johan, Sztuk-Dambietz, Jolanta, Tolstikova, Alexandra, Chapman, Henry N., Ros, Alexandra, Barty, Anton, Fromme, Petra, Mancuso, Adrian P., Schmidt, Marius, Cruz Villarreal, Jorvani, Yefanov, Oleksandr, Mariani, Valerio, White, Thomas A., and Kupitz, Christopher

Subjects
Enzyme substrate complex, Millisecond, Materials science, Bacterial antibiotic resistance, Chemical physics, Atomic resolution, Direct observation, Substrate (chemistry), Time resolution, ddc:610, Ultrashort pulse
Abstract

In this study, we follow the diffusion and buildup of occupancy of the substrate ceftriaxone inM. tuberculosisβ-lactamase BlaC microcrystals by structural analysis of the enzyme substrate complex at single millisecond time resolution. We also show the binding and the reaction of an inhibitor, sulbactam, on a slower millisecond time scale. We use the ‘mix-and-inject’ technique to initiate these reactions by diffusion, and determine the resulting structures by serial crystallography using ultrafast, intense X-ray pulses from the European XFEL (EuXFEL) arriving at MHz repetition rates. Here, we show how to use the EuXFEL pulse structure to dramatically increase the size of the data set and thereby the quality and time resolution of “molecular movies” which unravel ligand binding and enzymatically catalyzed reactions. This shows the great potential for the EuXFEL as a tool for biomedically relevant research, particularly, as shown here, for investigating bacterial antibiotic resistance.One Sentence SummaryDirect observation of fast ligand binding in a biomedically relevant enzyme at near atomic resolution with MHz X-ray pulses at the European XFEL.

Published
2020

46. Femtosecond X-ray protein nanocrystallography

Author

Chapman, Henry N., Fromme, Petra, Barty, Anton, White, Thomas A., Kirian, Richard A., Aquila, Andrew, Hunter, Mark S., Schulz, Joachim, DePonte, Daniel P., Weierstall, Uwe, Doak, Bruce R., Maia, Filipe R. N. C., Martin, Andrew V., Schlichting, Ilme, Lomb, Lukas, Coppola, Nicola, Shoeman, Robert L., Epp, Sascha W., Hartmann, Robert, Rolles, Daniel, Rudenko, Artem, Foucar, Lutz, Kimmel, Nils, Weidenspointner, Georg, Holl, Peter, Liang, Mengning, Barthelmess, Miriam, Caleman, Carl, Boutet, Sébastien, Bogan, Michael J., Krzywinski, Jacek, Bostedt, Christoph, Bajt, Saša, Gumprecht, Lars, Rudek, Benedikt, Erk, Benjamin, Schmidt, Carlo, Hömke, André, Reich, Christian, Pietschner, Daniel, Strüder, Lothar, Hauser, Günter, Gorke, Hubert, Ullrich, Joachim, Herrmann, Sven, Schaller, Gerhard, Schopper, Florian, Soltau, Heike, Kühnel, Kai-Uwe, Messerschmidt, Marc, Bozek, John D., Hau-Riege, Stefan P., Frank, Matthias, Hampton, Christina Y., Sierra, Raymond G., Starodub, Dmitri, Williams, Garth J., Hajdu, Janos, Timneanu, Nicusor, Seibert, Marvin M., Andreasson, Jakob, Rocker, Andrea, Jönsson, Olof, Svenda, Martin, Stern, Stephan, Nass, Karol, Andritschke, Robert, Schröter, Claus-Dieter, Krasniqi, Faton, Bott, Mario, Schmidt, Kevin E., Wang, Xiaoyu, Grotjohann, Ingo, Holton, James M., Barends, Thomas R. M., Neutze, Richard, Marchesini, Stefano, Fromme, Raimund, Schorb, Sebastian, Rupp, Daniela, Adolph, Marcus, Gorkhover, Tais, Andersson, Inger, Hirsemann, Helmut, Potdevin, Guillaume, Graafsma, Heinz, Nilsson, Björn, and Spence, John C. H.

Published
2011
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47. New Structural Insights into the Function of the Active Full Length Human Taspase1: A Novel Anticancer Therapeutic Target

Author

Jernigan, Rebecca J., primary, Nagaratnam, Nirupa, additional, Thifault, Darren, additional, Delker, Silvia, additional, Zacks, Michele, additional, Edwards, Thomas, additional, Sambucetti, Lidia, additional, Tong, Liang, additional, Fromme, Raimund, additional, Schneider, Joel, additional, Hsieh, James, additional, Mroczkowski, Barbra, additional, Flint, Andrew, additional, Fromme, Petra, additional, and Garcia, Jose M., additional

Published
2020
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48. Membrane Protein Dynamics Revealed by X-Ray Scattering with a Femtosecond Free-Electron Laser

Author

Grant, Thomas D., primary, Perera, Suchithranga M., additional, Salas-Estrada, Leslie A., additional, Struts, Andrey V., additional, Chawla, Udeep, additional, Xu, Xiaolin, additional, Fried, Steven D., additional, Weerasinghe, Nipuna, additional, Mendez, D., additional, Alvarez, R., additional, Karpos, K., additional, Lisova, S., additional, Zaare, S., additional, Nazari, R., additional, Zatsepsin, N.A., additional, Singharoy, Abhishek, additional, Boutet, S., additional, Carbajo, S., additional, Hunter, M.S., additional, Liang, M., additional, Seaberg, M.D., additional, Fromme, Raimund, additional, Fromme, Petra, additional, Grossfield, Alan, additional, Kirian, Richard A., additional, and Brown, Michael F., additional

Published
2020
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49. Rhodopsin's Ultra-Fast Activation Dynamics in Bilayer and Micelle Environments

Author

Salas-Estrada, Leslie A., primary, Grant, Thomas D., additional, Perera, Suchithranga M., additional, Struts, Andrey V., additional, Chawla, Udeep, additional, Xu, Xiaolin, additional, Fried, Steven D., additional, Weerasinghe, Nipuna, additional, Mendez, D., additional, Alvarez, R., additional, Karpos, K., additional, Lisova, S., additional, Zaare, S., additional, Nazari, R., additional, Zatsepsin, N.A., additional, Singharoy, Abhishek, additional, Boutet, S., additional, Carbajo, S., additional, Hunter, M.S., additional, Liang, M., additional, Seaberg, M.D., additional, Fromme, Raimund, additional, Fromme, Petra, additional, Kirian, Richard A., additional, Brown, Michael F., additional, and Grossfield, Alan, additional

Published
2020
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