Article
Biochemistry & Molecular Biology
Christopher J. Ohmer, Medhanjali Dasgupta, Anjali Patwardhan, Isabel Bogacz, Corey Kaminsky, Margaret D. Doyle, Percival Yang-Ting Chen, Stephen M. Keable, Hiroki Makita, Philipp S. Simon, Ramzi Massad, Thomas Fransson, Ruchira Chatterjee, Asmit Bhowmick, Daniel W. Paley, Nigel W. Moriarty, Aaron S. Brewster, Leland B. Gee, Roberto Alonso-Mori, Frank Moss, Franklin D. Fuller, Alexander Batyuk, Nicholas K. Sauter, Uwe Bergmann, Catherine L. Drennan, Vittal K. Yachandra, Junko Yano, Jan F. Kern, Stephen W. Ragsdale
Summary: Methyl-Coenzyme M Reductase (MCR) is a crucial enzyme in methanogenesis that has significant implications for global energy balances. However, the structure and reaction mechanism of this enzyme are still poorly understood.
JOURNAL OF INORGANIC BIOCHEMISTRY
(2022)
Article
Biochemical Research Methods
Deepshika Gilbile, Megan L. Shelby, Artem Y. Lyubimov, Jennifer L. Wierman, Diana C. F. Monteiro, Aina E. Cohen, Silvia Russi, Matthew A. Coleman, Matthias Frank, Tonya L. Kuhl
Summary: The development of versatile, inexpensive, and robust polymer microfluidic chips for serial X-ray crystallography allows for efficient sample delivery and measurements at both synchrotrons and X-ray free electron lasers. The chips offer advantages such as low sample consumption, minimal background scattering, and precise control over sample flow, making them suitable for a wide range of protein crystallography experiments.
Article
Multidisciplinary Sciences
Oliver F. Harder, Sarah V. Barrass, Marcel Drabbels, Ulrich J. Lorenz
Summary: Microsecond time-resolved cryo-EM allows for observation of fast protein dynamics, as demonstrated by the contraction of the CCMV capsid induced by a pH jump.
NATURE COMMUNICATIONS
(2023)
Article
Chemistry, Multidisciplinary
Hongjie Li, Yoshiki Nakajima, Takashi Nomura, Michihiro Sugahara, Shinichiro Yonekura, Siu Kit Chan, Takanori Nakane, Takahiro Yamane, Yasufumi Umena, Mamoru Suzuki, Tetsuya Masuda, Taiki Motomura, Hisashi Naitow, Yoshinori Matsuura, Tetsunari Kimura, Kensuke Tono, Shigeki Owada, Yasumasa Joti, Rie Tanaka, Eriko Nango, Fusamichi Akita, Minoru Kubo, So Iwata, Jian-Ren Shen, Michihiro Suga
Summary: The study successfully captured the structural dynamics of light-sensitive proteins during the S-1 to S-2 transition using pump-probe time-resolved serial femtosecond crystallography (TR-SFX), while also avoiding the influence of light contamination, revealing key structural changes important for the water-splitting reaction.
Review
Biophysics
Marcus Fischer
Summary: This review discusses practical aspects of preparing, acquiring, and analyzing X-ray crystallography data at room temperature, aiming to demystify preconceived impracticalities that freeze progress of routine room temperature data collection at synchrotron sources. Examples are provided as conceptual and experimental templates, illustrating the diversity and utility of gaining novel insights into protein conformational landscapes. An integrative view of protein conformational dynamics offers opportunities to advance basic and biomedical research.
QUARTERLY REVIEWS OF BIOPHYSICS
(2021)
Article
Chemistry, Multidisciplinary
Sylvain Aumonier, Sylvain Engilberge, Nicolas Caramello, David von Stetten, Guillaume Gotthard, Gordon A. Leonard, Christoph Mueller-Dieckmann, Antoine Royant
Summary: The development of serial crystallography has revitalized room-temperature macromolecular crystallography. This study describes the room-temperature structural study of a reaction intermediate using a synchrotron. The authors were able to monitor the relaxation process of the intermediate and discovered unexpected protein rearrangements.
Article
Biochemistry & Molecular Biology
Michael A. A. Hough, Filippo Prischi, Jonathan A. R. Worrall
Summary: The interaction between macromolecular proteins and small molecule ligands is crucial for cellular function, and studying this interaction is essential for understanding the structure and function of proteins and ligands.
FRONTIERS IN MOLECULAR BIOSCIENCES
(2023)
Review
Biophysics
Doeke R. Hekstra
Summary: Proteins play crucial roles in guiding the flow of information, energy, and matter in life by accelerating transport and chemical reactions, modulating these reactions, and forming dynamic supramolecular assemblies. Conformational change is fundamental to functional transitions, and time-resolved X-ray diffraction methods have been successful in characterizing these transitions. This review focuses on emerging techniques that investigate the dynamic basis of function in proteins without native light-dependent transitions, and discusses potential extensions and future possibilities. Additionally, the review examines the challenges posed by the weaker and more distributed signals in these data, pushing the limits of analytical methods. Overall, these new methods are establishing a powerful paradigm for studying the physics of protein function.
ANNUAL REVIEW OF BIOPHYSICS
(2023)
Article
Biology
Tamar Skaist Mehlman, Justin T. Biel, Syeda Maryam Azeem, Elliot R. Nelson, Sakib Hossain, Louise Dunnett, Neil G. Paterson, Alice Douangamath, Romain Talon, Danny Axford, Helen Orins, Frank von Delft, Daniel A. Keedy, Qiang Cui
Summary: Much of our understanding of protein-ligand interactions comes from X-ray crystal structures determined at cryogenic temperature. However, the impact of room-temperature crystallography on protein-ligand complexes is not well understood. Our study shows that at room temperature, fewer ligands bind and often with weaker affinity, but with temperature-dependent differences in binding poses, solvation, binding sites, and protein conformational responses. This work suggests that existing cryo-temperature structures may provide an incomplete picture and highlights the potential of room-temperature crystallography in revealing distinct conformational modes of protein-ligand systems.
Article
Biochemical Research Methods
Zhongrui Liu, Kevin K. Gu, Megan L. Shelby, Deepshika Gilbile, Artem Y. Lyubimov, Silvia Russi, Aina E. Cohen, Sankar Raju Narayanasamy, Sabine Botha, Christopher Kupitz, Raymond G. Sierra, Fredric Poitevin, Antonio Gilardi, Stella Lisova, Matthew A. Coleman, Matthias Frank, Tonya L. Kuhl
Summary: Over the past two decades, serial X-ray crystallography has made significant progress in determining the structures of proteins. This study presents an improved design of a polymer microfluidic chip that facilitates the delivery of fragile and small crystals to X-ray beams. The chip is customizable for different types of crystals and offers a user-friendly, cost-effective, and robust sample-delivery platform. Experimental results demonstrate the effectiveness of the chip design in obtaining high-resolution data sets for various proteins. The improved chip design provides a powerful sample-delivery tool for structural biologists, especially when working with limited sample volumes and fragile crystals.
ACTA CRYSTALLOGRAPHICA SECTION D-STRUCTURAL BIOLOGY
(2023)
Article
Physics, Multidisciplinary
Can B. Uzundal, Sasawat Jamnuch, Emma Berger, Clarisse Woodahl, Paul Manset, Yasuyuki Hirata, Toshihide Sumi, Angelique Amado, Hisazumi Akai, Yuya Kubota, Shigeki Owada, Kensuke Tono, Makina Yabashi, John W. Freeland, Craig P. Schwartz, Walter S. Drisdell, Iwao Matsuda, Tod A. Pascal, Alfred Zong, Michael Zuerch
Summary: Second harmonic generation (SHG) spectroscopy is commonly used to investigate surface chemistry, interfacial chemistry, and symmetry properties in solids, but the complexity of novel materials makes interpretation of experiments difficult. Polarization-resolved extreme ultraviolet (XUV-SHG) spectroscopy demonstrates element-specific angular anisotropy investigations for the first time, paving the way for future angle and time-resolved XUV-SHG studies with elemental specificity in condensed matter systems.
PHYSICAL REVIEW LETTERS
(2021)
Article
Chemistry, Multidisciplinary
Jangwoo Kim, Ki Hyun Nam
Summary: When X-rays pass through a material, heat is generated and transferred, which can affect the temperature of the sample and the experimental environment in serial crystallography experiments. This study investigated radiation damage and measured the level of heating near the X-ray interaction point. The findings provide valuable information for obtaining more accurate experimental parameters.
APPLIED SCIENCES-BASEL
(2023)
Editorial Material
Crystallography
Ki Hyun Nam
Summary: Serial crystallography (SX) is an emerging technique to determine macromolecules at room temperature. It provides time-resolved dynamics of target molecules through pump-probe experiments. SX has rapidly developed as a technique for studying room-temperature structures and molecular dynamics of biomolecules. The development of sample delivery techniques and data processing programs is accelerating SX research.
Article
Biochemical Research Methods
Jack B. Greisman, Kevin M. Dalton, Candice J. Sheehan, Margaret A. Klureza, Igor Kurinov, Doeke R. Hekstra
Summary: Single-wavelength anomalous diffraction (SAD) is a common method for solving the phase problem in macromolecular structures. This study presents a strategy to obtain high-quality data from room-temperature, single-crystal experiments, and successfully applies it to solve four structures of three model systems. The resulting data sets allow for automatic phasing and model building, and reveal alternate conformations of proteins at room temperature.
ACTA CRYSTALLOGRAPHICA SECTION D-STRUCTURAL BIOLOGY
(2022)
Article
Biochemistry & Molecular Biology
Valeria Conforte, Lisandro Horacio Otero, Laila Toum, Serena Sirigu, Giuliano Tomas Antelo, Jimena Rinaldi, Sabrina Foscaldi, Sebastian Klinke, Leonard Michel Gabriel Chavas, Adrian Alberto Vojnov, Fernando Alberto Goldbaum, Florencia Malamud, Hernan Ruy Bonomi
Summary: The study reveals that variants affecting the XccBphP photocycle have an impact on xanthan production and stomatal aperture in the bacterial signaling pathway. These findings suggest a novel approach where the Pr state of XccBphP acts as a negative regulator of virulence mechanisms in Xcc.
Article
Biochemistry & Molecular Biology
Celine Lafaye, Sylvain Aumonier, Joaquim Torra, Luca Signor, David von Stetten, Marjolaine Noirclerc-Savoye, Xiaokun Shu, Ruben Ruiz-Gonzalez, Guillaume Gotthard, Antoine Royant, Santi Nonell
Summary: The study shows that modifying the chromophore structure of miniSOG can significantly improve its photosensitizing properties. Replacing flavin mononucleotide (FMN) with riboflavin (RF) without the bulky phosphate group increases the yield of singlet oxygen. Mutagenizing residues stabilizing the phosphate group allows for the modification of miniSOG's photophysical properties.
PHOTOCHEMICAL & PHOTOBIOLOGICAL SCIENCES
(2022)
Article
Chemistry, Multidisciplinary
Lea Albert, Jatin Nagpal, Wieland Steinchen, Lei Zhang, Laura Werel, Nemanja Djokovic, Dusan Ruzic, Malte Hoffarth, Jing Xu, Johanna Kaspareit, Frank Abendroth, Antoine Royant, Gert Bange, Katarina Nikolic, Soojin Ryu, Yali Dou, Lars-Oliver Essen, Olalla Vazquez
Summary: A genuine on-off optochemical probe has been developed for hematopoietic control in zebrafish with unprecedented stability. This interdisciplinary study uncovers the mechanism of action of the probe, providing a new method for future investigations.
ACS CENTRAL SCIENCE
(2022)
Article
Biochemistry & Molecular Biology
Matthew J. Rodrigues, Nitai Giri, Antoine Royant, Yang Zhang, Rachel Bolton, Gwyndaf Evans, Steve E. Ealick, Tadhg Begley, Ivo Tews
Summary: The Pdx1 enzyme catalyzes the formation of pyridoxal 5-phosphate (PLP) through an imine relay mechanism, involving stereoselective deprotonation and substrate-assisted phosphate elimination. The I-333 intermediate, characterized by structural, UV-vis absorption spectroscopy and mass spectrometry analyses, plays a central role in PLP biosynthesis.
RSC CHEMICAL BIOLOGY
(2022)
Article
Chemistry, Multidisciplinary
Manuel Maestre-Reyna, Cheng-Han Yang, Eriko Nango, Wei-Cheng Huang, Eka Putra Gusti Ngurah Putu, Wen-Jin Wu, Po-Hsun Wang, Sophie Franz-Badur, Martin Saft, Hans-Joachim Emmerich, Hsiang-Yi Wu, Cheng-Chung Lee, Kai-Fa Huang, Yao-Kai Chang, Jiahn-Haur Liao, Jui-Hung Weng, Wael Gad, Chiung-Wen Chang, Allan H. Pang, Michihiro Sugahara, Shigeki Owada, Yuhei Hosokawa, Yasumasa Joti, Ayumi Yamashita, Rie Tanaka, Tomoyuki Tanaka, Fangjia Luo, Kensuke Tono, Kai-Cheng Hsu, Stephan Kiontke, Igor Schapiro, Roberta Spadaccini, Antoine Royant, Junpei Yamamoto, So Iwata, Lars-Oliver Essen, Yoshitaka Bessho, Ming-Daw Tsai
Summary: In this study, time-resolved serial femtosecond X-ray crystallography was used to analyze the entanglement between FAD and its redox sensor triad. It was found that this process regulates FAD re-hybridization and protonation.
Article
Multidisciplinary Sciences
Sandra Mous, Guillaume Gotthard, David Ehrenberg, Saumik Sen, Tobias Weinert, Philip J. M. Johnson, Daniel James, Karol Nass, Antonia Furrer, Demet Kekilli, Pikyee Ma, Steffen Bruenle, Cecilia Maria Casadei, Isabelle Martiel, Florian Dworkowski, Dardan Gashi, Petr Skopintsev, Maximilian Wranik, Gregor Knopp, Ezequiel Panepucci, Valerie Panneels, Claudio Cirelli, Dmitry Ozerov, Gebhard F. X. Schertler, Meitian Wang, Chris Milne, Joerg Standfuss, Igor Schapiro, Joachim Heberle, Przemyslaw Nogly
Summary: By combining time-resolved crystallography, spectroscopy, and multiscale simulations, we have elucidated the molecular mechanism and structural dynamics of a chloride-pumping rhodopsin throughout the transport cycle. We identified transient anion-binding sites, provided evidence for the use of light energy in the pumping mechanism, and discovered molecular gates that ensure unidirectional transport. These findings reveal key mechanistic features that enable finely controlled chloride transport across the cell membrane in this light-powered chloride ion pump.
Article
Chemistry, Multidisciplinary
Sylvain Aumonier, Sylvain Engilberge, Nicolas Caramello, David von Stetten, Guillaume Gotthard, Gordon A. Leonard, Christoph Mueller-Dieckmann, Antoine Royant
Summary: The development of serial crystallography has revitalized room-temperature macromolecular crystallography. This study describes the room-temperature structural study of a reaction intermediate using a synchrotron. The authors were able to monitor the relaxation process of the intermediate and discovered unexpected protein rearrangements.
Article
Multidisciplinary Sciences
Thomas Gruhl, Tobias Weinert, Matthew J. Rodrigues, Christopher J. Milne, Giorgia Ortolani, Karol Nass, Eriko Nango, Saumik Sen, Philip J. M. Johnson, Claudio Cirelli, Antonia Furrer, Sandra Mous, Petr Skopintsev, Daniel James, Florian Dworkowski, Petra Bath, Demet Kekilli, Dmitry Ozerov, Rie Tanaka, Hannah Glover, Camila Bacellar, Steffen Brunle, Cecilia M. Casadei, Azeglio D. Diethelm, Dardan Gashi, Guillaume Gotthard, Ramon Guixa-Gonzalez, Yasumasa Joti, Victoria Kabanova, Gregor Knopp, Elena Lesca, Pikyee Ma, Isabelle Martiel, Jonas Muhle, Shigeki Owada, Filip Pamula, Daniel Sarabi, Oliver Tejero, Ching-Ju Tsai, Niranjan Varma, Anna Wach, Sebastien Boutet, Kensuke Tono, Przemyslaw Nogly, Xavier Deupi, So Iwata, Richard Neutze, Jorg Standfuss, Gebhard Schertler, Valerie Panneels
Summary: This article reveals the early stages of the vision process through ultrafast time-resolved crystallography experiments, providing the latest research on vertebrate vision and insights into the molecular mechanisms of agonist-mediated GPCR activation.
Article
Biochemical Research Methods
Theodorus W. J. Gadella, Laura van Weeren, Jente Stouthamer, Mark A. Hink, Anouk H. G. Wolters, Ben N. G. Giepmans, Sylvain Aumonier, Jerome Dupuy, Antoine Royant
Summary: We have evolved mScarlet3, a cysteine-free monomeric red fluorescent protein, with fast and complete maturation, high brightness, quantum yield (75%) and fluorescence lifetime (4.0 ns). The crystal structure of mScarlet3 reveals a barrel rigidified at one of its heads by a large hydrophobic patch of internal residues. mScarlet3 performs well as a fusion tag, shows no apparent cytotoxicity, and outperforms existing red fluorescent proteins as a Forster resonance energy transfer acceptor and as a reporter in transient expression systems.
Article
Multidisciplinary Sciences
Pedram Mehrabi, Sihyun Sung, David von Stetten, Andreas Prester, Caitlin E. Hatton, Stephan Kleine-Doepke, Alexander Berkes, Gargi Gore, Jan-Philipp Leimkohl, Hendrik Schikora, Martin Kollewe, Holger Rohde, Matthias Wilmanns, Friedjof Tellkamp, Eike C. Schulz
Summary: The authors introduce the spitrobot, a crystal plunger, enabling cryo-trapping with millisecond time-resolution via the liquid application method (LAMA). Ligand binding and reaction intermediates are demonstrated in three different enzymes. We also trap reaction intermediates and conformational changes in macroscopic crystals of tryptophan synthase to demonstrate that the spitrobot enables insight into catalytic events.
NATURE COMMUNICATIONS
(2023)
Article
Instruments & Instrumentation
Kate Mary Louise Smith, Ezequiel Panepucci, Jakub Wojciech Kaminski, Sylvain Aumonier, Chia-Yiang Huang, Deniz Eris, Dominik Buntschu, Nathalie Meier, Wayne Glettig, Katherine Evelyn McAuley, Meitian Wang, May Elizabeth Sharpe, Justyna Aleksandra Wojdyla
Summary: Recent advances in automation have enabled the development of unattended data collection services in several synchrotron facilities globally. This paper focuses on the Swiss Light Source, where the implementation of new high-throughput sample changers and the commissioning of the Fast Fragment and Compound Screening pipeline provided an opportunity for automated data acquisition. The paper discusses the upgrades in the DA+ microservice software stack, the automatic loop-centering service, and the deployment of the Smart Digital User (SDU) software for unattended data collection. The SDU software plays a crucial role in communication between services, ensuring sample and device safety, sample centering, alignment with grid-based X-ray diffraction, and ultimately, data collection.
JOURNAL OF SYNCHROTRON RADIATION
(2023)
Article
Biochemistry & Molecular Biology
Kirill Kovalev, Fedor Tsybrov, Alexey Alekseev, Vitaly Shevchenko, Dmytro Soloviov, Sergey Siletsky, Gleb Bourenkov, Michael Agthe, Marina Nikolova, David von Stetten, Roman Astashkin, Sergey Bukhdruker, Igor Chizhov, Antoine Royant, Alexander Kuzmin, Ivan Gushchin, Riccardo Rosselli, Francisco Rodriguez-Valera, Nikolay Ilyinskiy, Andrey Rogachev, Valentin Borshchevskiy, Thomas R. Schneider, Ernst Bamberg, Valentin Gordeliy
Summary: By studying a bacterial inward proton-pumping rhodopsin, researchers have uncovered its molecular mechanism and revealed how proton wires mediate ion selectivity and proton transport through cell membrane. Proton transport is essential for cell life and understanding the mechanisms behind it is a challenge. The study presents a comprehensive function-structure analysis of a light-driven bacterial inward proton pump, providing insights into proton translocation and the use of synchrotron source for rhodopsin studies. The results also have implications for optogenetics.
NATURE STRUCTURAL & MOLECULAR BIOLOGY
(2023)
Review
Crystallography
Sam Horrell, Gianluca Santoni, Andrea Thorn
Summary: This review summarizes the structural and functional information of SARS-CoV-2 nsp15, discusses structure-based drug design efforts and complementary knowledge related to other coronaviruses, providing a clear starting point for researchers interested in studying nsp15 as a novel drug target for treating COVID-19.
CRYSTALLOGRAPHY REVIEWS
(2022)
Article
Biochemistry & Molecular Biology
Landon Zarowny, Damien Clavel, Ryan Johannson, Kevin Duarte, Hadrien Depernet, Jerome Dupuy, Heather Baker, Alex Brown, Antoine Royant, Robert E. Campbell
Summary: In this study, we successfully created a new fluorescent protein, NeonCyan1, and two additional mutants with distinct spectral hues through modification of mNeonGreen. Structural, computational, and photophysical characterization of these proteins provided insight into the factors that control the fluorescence emission color. Although not recommended as replacements for contemporary fluorescent proteins, these new variants demonstrate potential suitability for live cell imaging applications.
PROTEIN ENGINEERING DESIGN & SELECTION
(2022)
