Article
Multidisciplinary Sciences
Amani A. Hariri, Sharon S. Newman, Steven Tan, Dan Mamerow, Alexandra M. Adams, Nicolo Maganzini, Brian L. Zhong, Michael Eisenstein, Alexander R. Dunn, H. Tom Soh
Summary: One major challenge in enzyme-linked immunosorbent assays (ELISAs) is distinguishing true signal from non-specific binding. The authors developed a Single-Molecule Colocalization Assay (SiMCA) that eliminates this issue and allows for reproducible detection of picomolar protein concentrations.
NATURE COMMUNICATIONS
(2022)
Article
Chemistry, Multidisciplinary
Simanta Sarani Paul, Aaron Lyons, Russell Kirchner, Michael T. Woodside
Summary: Protein aggregation is a characteristic of many neurodegenerative diseases. This study used single-molecule mass photometry to track the populations of different oligomers during aggregation of tau protein. The results revealed an equilibrium between monomers, dimers, and trimers before aggregation, and a continuous decrease in monomer population accompanied by an increase in smaller oligomers and subsequent incorporation into larger oligomers and fibrils during aggregation.
Article
Nanoscience & Nanotechnology
Jan Becker, Jack S. Peters, Ivor Crooks, Seham Helmi, Marie Synakewicz, Benjamin Schuler, Philipp Kukura
Summary: Label-free detection of single biomolecules in solution has been achieved using various experimental approaches, but our understanding of the optical contrast and its relationship with atomic structure, as well as the measurement sensitivity and precision, is still unclear. In this study, a Fourier optics approach combined with an atomic structure-based molecular polarizability model is used to simulate mass photometry experiments, resulting in excellent agreement with experimentally determined parameters. This provides a general framework for light-based single-molecule detection and quantification that is independent of the optical detection approach chosen.
Article
Chemistry, Multidisciplinary
Serena Monaco, Jesus Angulo, Matthew Wallace
Summary: We developed a new tool called imaging STD NMR by combining saturation transfer difference NMR (STDNMR) with chemical shift imaging (CSI) and controlled concentration gradients of small molecule ligands. This method allows for the assessment of protein-ligand interactions, determination of dissociation constants (K(D)), and evaluation of binding specificity in a single NMR tube, saving time compared to traditional titration methods. Imaging STD NMR also allows for screening of non-specific binders, making it a potential tool to speed up and optimize the drug discovery process.
JOURNAL OF THE AMERICAN CHEMICAL SOCIETY
(2023)
Review
Biology
M. Shahid Mansuri, Kenneth Williams, Angus C. Nairn
Summary: Recent technological advances have enabled single-cell proteomics to unravel the role of protein expression, post-translational modifications, and protein interactions in determining cell state in both health and disease.
COMMUNICATIONS BIOLOGY
(2023)
Article
Nanoscience & Nanotechnology
Carlos J. Bueno-Alejo, Marina Santana Vega, Amanda K. Chaplin, Chloe Farrow, Alexander Axer, Glenn A. Burley, Cyril Dominguez, Hesna Kara, Vasileios Paschalis, Sumera Tubasum, Ian C. Eperon, Alasdair W. Clark, Andrew J. Hudson
Summary: Single-molecule imaging is important for studying the behavior and interactions of biological molecules. However, the irreversible adsorption of components onto surfaces poses a challenge for precise sampling. Interferometric scattering microscopy (iSCAT) can detect the adsorption and desorption of unlabeled proteins on glass surfaces. A surface passivation method using high-density perfluoroalkane brushes allows for continuous monitoring of protein adsorption and desorption without irreversible binding.
ACS APPLIED MATERIALS & INTERFACES
(2022)
Article
Chemistry, Multidisciplinary
Ting Xue, Weikun Wu, Ning Guo, Chengyong Wu, Jian Huang, Lipeng Lai, Hong Liu, Yalun Li, Tianyuan Wang, Yuxi Wang
Summary: Research shows that the RBD of the SARS-CoV-2 virus S protein plays a key role in viral cell attachment, and mutations may affect the potency of vaccines and antibodies. Using in silico methods, the study predicted the impact of RBD mutations on its binding affinity to cell receptors, with experimental results showing agreement with the predictions. This demonstrates the potential of in silico tools in forecasting more powerful virus mutants to aid in the development of broadly neutralizing vaccines and antibodies.
Article
Oncology
Keelan Z. Guiley, Kevan M. Shokat
Summary: The transcription factor and tumor suppressor protein p53 is frequently mutated and inactivated in cancer. This study successfully identified covalent compounds that selectively react with the p53 somatic mutant cysteine Y220C and restore wild-type thermal stability.
Article
Multidisciplinary Sciences
Di Wu, Grzegorz Piszczek
Summary: Mass photometry (MP) is a label- and immobilization-free technique that allows for the precise determination of antigen-antibody interaction affinities and provides information on the stoichiometry and oligomeric state of the proteins. It is a simple and straightforward method that requires minimal quantities of protein and no expensive consumables, making it ideal for studying protein-protein interactions.
JOVE-JOURNAL OF VISUALIZED EXPERIMENTS
(2021)
Article
Multidisciplinary Sciences
Jesper Levring, Daniel S. Terry, Zeliha Kilic, Gabriel Fitzgerald, Scott Blanchard, Jue Chen
Summary: The cystic fibrosis transmembrane conductance regulator (CFTR) is an anion channel that regulates salt and fluid homeostasis across epithelial membranes. Alterations in CFTR cause cystic fibrosis, a fatal disease without a cure. In this study, the authors investigate the structure and function of CFTR, specifically focusing on the dimerization of its nucleotide-binding domains (NBDs) and the allosteric gating mechanism that regulates chloride conductance. They also explore the effects of disease-causing substitutions on NBD dimerization and propose implications for potential clinical therapies.
Article
Chemistry, Analytical
Clarice E. Froehlich, Jiayi He, Christy L. Haynes
Summary: Investigating the interactions between small, charged molecules and aptamers using surface plasmon resonance (SPR) is challenging due to the low response of small molecules and nonspecific electrostatic interactions. However, aptamers are increasingly used in sensors for small molecule detection. This study develops a method to characterize the interaction between the charged small molecule serotonin and an aptamer using streptavidin-biotin capture and a high-ionic-strength buffer. These techniques provide insight into working with small molecules and offer the potential for improving SPR sensors.
ANALYTICAL CHEMISTRY
(2023)
Article
Chemistry, Analytical
Matteo Scalabrin, Matteo Nadai, Martina Tassinari, Sara Lago, Filippo Doria, Ilaria Frasson, Mauro Freccero, Sara N. Richter
Summary: This study developed a rapid method to screen for single G4 ligands and successfully identified nonplanar pyrazolopyrimidines that selectively recognize and stabilize HIV-1 G4. These compounds inhibit HIV-1 replication, potentially providing a new approach for the development of novel G4-based therapeutics.
ANALYTICAL CHEMISTRY
(2021)
Review
Biochemistry & Molecular Biology
Byungju Kim, Jincheol Seol, Yoon Ki Kim, Jong-Bong Lee
Summary: Single-molecule fluorescence imaging allows for the observation of circular mRNA formation and the intricate interplay between messenger RNA and ribosomal proteins during translation. This detailed visualization method could potentially enable direct manipulation of interactions within mRNA-protein complexes.
EXPERIMENTAL AND MOLECULAR MEDICINE
(2023)
Article
Physics, Applied
Houman Mirzaalian Dastjerdi, Mahyar Dahmardeh, Andre Gemeinhardt, Reza Gholami Mahmoodabadi, Harald Koestler, Vahid Sandoghdar
Summary: This article discusses the sensitivity of interferometric detection of Rayleigh scattering (iSCAT) and emphasizes the importance of data analysis strategies and algorithms. The impact of speckle-like background and technical issues on the signal-to-noise ratio is discussed.
JOURNAL OF PHYSICS D-APPLIED PHYSICS
(2022)
Article
Chemistry, Multidisciplinary
Sean A. Burnap, Weston B. Struwe
Summary: This study demonstrates how proline substitutions used for spike stabilization in SARS-CoV-2 vaccine design directly impact ACE2 receptor interactions. Conformational changes and ACE2 binding are influenced by spike variants and temperature, but are independent of site-specific N-glycosylation.
CHEMICAL COMMUNICATIONS
(2022)
Article
Biophysics
Lewis S. Mosby, Nikolas Hundt, Gavin Young, Adam Fineberg, Marco Polin, Satyajit Mayor, Philipp Kukura, Darius Koster
BIOPHYSICAL JOURNAL
(2020)
Article
Chemistry, Physical
Raj Pandya, Richard Y. S. Chen, Qifei Gu, Jeffrey Gorman, Florian Auras, Jooyoung Sung, Richard Friend, Philipp Kukura, Christoph Schnedermann, Akshay Rao
JOURNAL OF PHYSICAL CHEMISTRY A
(2020)
Article
Chemistry, Multidisciplinary
Maria A. Lebedeva, Elena Palmieri, Philipp Kukura, Stephen P. Fletcher
Article
Biochemistry & Molecular Biology
Yiwen Li, Weston B. Struwe, Philipp Kukura
NUCLEIC ACIDS RESEARCH
(2020)
Review
Chemistry, Multidisciplinary
Lee Priest, Jack S. Peters, Philipp Kukura
Summary: Our ability to detect, image, and quantify nanoscopic objects and molecules with visible light has dramatically improved in recent decades, with recent developments in light scattering reaching single-molecule sensitivity. These developments have the advantages of universal applicability and the ability to obtain more information about the species of interest.
Article
Biochemical Research Methods
Eric D. B. Foley, Manish S. Kushwah, Gavin Young, Philipp Kukura
Summary: Dynamic mass photometry allows label-free tracking and mass measurement of individual membrane-associated proteins diffusing on supported lipid bilayers. The approach can be used to monitor dynamic (dis)assembly of protein complexes.
Article
Nanoscience & Nanotechnology
Xuanhui Meng, Adar Sonn-Segev, Anne Schumacher, Daniel Cole, Gavin Young, Stephen Thorpe, Robert W. Style, Eric R. Dufresne, Philipp Kukura
Summary: Single particle tracking has broad applications in life and physical sciences, allowing observation and characterization of nano- and microscopic motion. This study demonstrates that micromirror-based total internal reflection dark field microscopy achieves efficient background suppression, with nanometer localization precision and high temporal resolution.
Review
Biochemistry & Molecular Biology
Roi Asor, Philipp Kukura
Summary: This article reviews recent advances in characterizing biomolecular structure, interactions, and dynamics using mass photometry (MP), which enables label-free detection and mass measurement of individual biomolecules in solution. Molecular counting and identification provide information on relative abundance and affinities, while dynamics offer insights into on- and off-rates. The molecular resolution of MP allows these measurements to be performed as a function of stoichiometry and assembly at equilibrium, which is different from many existing solution-based methods. With further improvements in assays and technological performance, MP is expected to provide mechanistic details of complex biomolecular processes.
CURRENT OPINION IN CHEMICAL BIOLOGY
(2022)
Article
Multidisciplinary Sciences
Nikolas Hundt, Daniel Cole, Max F. Hantke, Jack J. Miller, Weston B. Struwe, Philipp Kukura
Summary: This study uses mass photometry to investigate the polymerization process of actin and finds that the traditional nucleation-based models cannot explain the observed distribution of actin oligomers. Instead, the key step of filament formation is a slow transition between different states of an actin filament mediated by cation exchange or ATP hydrolysis. These findings have important implications for understanding the mechanism of actin nucleation and studying protein assembly at the molecular level.
Article
Chemistry, Multidisciplinary
Tayyaba Younas, Chang Liu, Weston B. Struwe, Philipp Kukura, Lizhong He
Summary: Researchers have engineered light-responsive RNA-protein nanowires that exhibit non-covalent light-switching ability. By utilizing the high affinity between RNA and protein, they replaced green fluorescence protein with an optogenetic protein pair system, allowing blue light-controlled photo-switching. These RNA-protein structures can serve as biocompatible frameworks for incorporating versatile elements such as RNA, DNA, and enzymes.
Article
Nanoscience & Nanotechnology
Jan Becker, Jack S. Peters, Ivor Crooks, Seham Helmi, Marie Synakewicz, Benjamin Schuler, Philipp Kukura
Summary: Label-free detection of single biomolecules in solution has been achieved using various experimental approaches, but our understanding of the optical contrast and its relationship with atomic structure, as well as the measurement sensitivity and precision, is still unclear. In this study, a Fourier optics approach combined with an atomic structure-based molecular polarizability model is used to simulate mass photometry experiments, resulting in excellent agreement with experimentally determined parameters. This provides a general framework for light-based single-molecule detection and quantification that is independent of the optical detection approach chosen.
Article
Chemistry, Physical
Fabian Soltermann, Weston B. Struwe, Philipp Kukura
Summary: This paper reviews recent advances in optical technologies for label-free in vitro measurements of protein-protein interactions, including an overview and comparison of existing techniques, as well as discussions on their advantages, limitations, and recent applications.
PHYSICAL CHEMISTRY CHEMICAL PHYSICS
(2021)
Article
Chemistry, Multidisciplinary
Xuanhui Meng, Philipp Kukura, Sanli Faez
Summary: This study presents a molecular force sensor and charge detector based on tracking tethered double-stranded DNA functionalised with charged nanoparticles. The approach allows for real-time detection and quantification of electrophoretic force and changes in particle charge state down to the sub-piconewton scale, providing an alternative route for studying structural and charge dynamics at the single molecule level.
Article
Biochemistry & Molecular Biology
Ankita Chadda, Alexander G. Kozlov, Binh Nguyen, Timothy M. Lohman, Eric A. Galburt
Summary: In this study, it was found that the DNA damage response in Mycobacterium tuberculosis differs from well-studied model bacteria. The DNA repair helicase UvrD1 in Mtb is activated through a redox-dependent process and is closely associated with the homo-dimeric Ku protein. Additionally, Ku protein is shown to stimulate the helicase activity of UvrD1.
JOURNAL OF MOLECULAR BIOLOGY
(2024)
