Publications by authors named "Martha E Brennich"

7 Publications

  • Page 1 of 1

Molecular model of a sensor of two-component signaling system.

Sci Rep 2021 May 24;11(1):10774. Epub 2021 May 24.

Research Center for Molecular Mechanisms of Aging and Age-Related Diseases, Moscow Institute of Physics and Technology, 141700, Dolgoprudny, Russia.

Two-component systems (TCS) are widespread signaling systems present in all domains of life. TCS typically consist of a signal receptor/transducer and a response regulator. The receptors (histidine kinases, chemoreceptors and photoreceptors) are often embedded in the membrane and have a similar modular structure. Chemoreceptors were shown to function in highly ordered arrays, with trimers of dimers being the smallest functional unit. However, much less is known about photoreceptors. Here, we use small-angle scattering (SAS) to show that detergent-solubilized sensory rhodopsin II in complex with its cognate transducer forms dimers at low salt concentration, which associate into trimers of dimers at higher buffer molarities. We then fit an atomistic model of the whole complex into the SAS data. The obtained results suggest that the trimer of dimers is "tripod"-shaped and that the contacts between the dimers occur only through their cytoplasmic regions, whereas the transmembrane regions remain unconnected.
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http://dx.doi.org/10.1038/s41598-021-89613-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8144572PMC
May 2021

Mutation-induced alterations of intra-filament subunit organization in vimentin filaments revealed by SAXS.

Soft Matter 2019 Feb;15(9):1999-2008

Institute for X-ray Physics, University of Goettingen, Friedrich-Hund-Platz 1, 37077 Göttingen, Germany.

Vimentin intermediate filaments constitute a distinct filament system in mesenchymal cells that is instrumental for cellular mechanics and migration. In vitro, the rod-like monomers assemble in a multi-step, salt-dependent manner into micrometer long biopolymers. To disclose the underlying mechanisms further, we employed small angle X-ray scattering on two recombinant vimentin variants, whose assembly departs at strategic points from the normal assembly route: (i) vimentin with a tyrosine to leucine change at position 117; (ii) vimentin missing the non-α-helical carboxyl-terminal domain. Y117L vimentin assembles into unit-length filaments (ULFs) only, whereas ΔT vimentin assembles into filaments containing a higher number of tetramers per cross section than normal vimentin filaments. We show that the shape and inner structure of these mutant filaments is significantly altered. ULFs assembled from Y117L vimentin contain more, less tightly bundled vimentin tetramers, and ΔT vimentin filaments preserve the number density despite the higher number of tetramers per filament cross-section.
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http://dx.doi.org/10.1039/c8sm02281jDOI Listing
February 2019

Smaller capillaries improve the small-angle X-ray scattering signal and sample consumption for biomacromolecular solutions.

J Synchrotron Radiat 2018 Jul 26;25(Pt 4):1113-1122. Epub 2018 Jun 26.

European Molecular Biology Laboratory (EMBL), Hamburg Outstation c/o DESY, Notkestrasse 85, 22607 Hamburg, Germany.

Radiation damage by intense X-ray beams at modern synchrotron facilities is one of the major complications for biological small-angle X-ray scattering (SAXS) investigations of macromolecules in solution. To limit the damage, samples are typically measured under a laminar flow through a cell (typically a capillary) such that fresh solution is continuously exposed to the beam during measurement. The diameter of the capillary that optimizes the scattering-to-absorption ratio at a given X-ray wavelength can be calculated a priori based on fundamental physical properties. However, these well established scattering and absorption principles do not take into account the radiation susceptibility of the sample or the often very limited amounts of precious biological material available for an experiment. Here it is shown that, for biological solution SAXS, capillaries with smaller diameters than those calculated from simple scattering/absorption criteria allow for a better utilization of the available volumes of radiation-sensitive samples. This is demonstrated by comparing two capillary diameters d (d = 1.7 mm, close to optimal for 10 keV; and d = 0.9 mm, which is nominally sub-optimal) applied to study different protein solutions at various flow rates. The use of the smaller capillaries ultimately allows one to collect higher-quality SAXS data from the limited amounts of purified biological macromolecules.
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http://dx.doi.org/10.1107/S1600577518007907DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6038601PMC
July 2018

Online Size-exclusion and Ion-exchange Chromatography on a SAXS Beamline.

J Vis Exp 2017 01 5(119). Epub 2017 Jan 5.

Groupe de Microscopie Electronique et Méthodes, Institut de Biologie Structurale;

Biological small angle X-ray scattering (BioSAXS) is a powerful technique in molecular and structural biology used to determine solution structure, particle size and shape, and surface-to-volume ratio of macromolecules. The technique is applicable to a very wide variety of solution conditions spanning a broad range of concentrations, pH values, ionic strengths, temperatures, additives, etc., but the sample is required to be monodisperse. This caveat led to the implementation of liquid chromatography systems on SAXS beamlines. Here, we describe the upstream integration of size-exclusion (SEC) and ion-exchange chromatography (IEC) on a beamline, different methods for optimal background subtraction, and data reduction. As an example, we describe how we use SEC- and IEC-SAXS on a fragment of the essential vaccinia virus protein D5, consisting of a D5N helicase domain. We determine its overall shape and molecular weight, showing the hexameric structure of the protein.
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http://dx.doi.org/10.3791/54861DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5409194PMC
January 2017

The filament forming reactions of vimentin tetramers studied in a serial-inlet microflow device by small angle x-ray scattering.

Biomicrofluidics 2016 Mar 16;10(2):024108. Epub 2016 Mar 16.

Institute for X-Ray Physics, Georg-August-Universität Göttingen , 37077 Göttingen, Germany.

The structural organization of metazoan cells and their shape are established through the coordinated interaction of a composite network consisting of three individual filament systems, collectively termed the cytoskeleton. Specifically, microtubules and actin filaments, which assemble from monomeric globular proteins, provide polar structures that serve motor proteins as tracks. In contrast, intermediate filaments (IFs) assemble from highly charged, extended coiled coils in a hierarchical assembly mechanism of lateral and longitudinal interaction steps into non-polar structures. IF proteins are expressed in a distinctly tissue-specific way and thereby serve to generate the precise plasticity of the respective cells and tissues. Accordingly, in the cell, numerous parameters such as pH and salt concentration are adjusted such that the generation of functional networks is ensured. Here, we transfer the problem for the mesenchymal IF protein vimentin to an in vitro setting and combine small angle x-ray scattering with microfluidics and finite element method simulations. Our approach is adapted to resolve the early assembly steps, which take place in the sub-second to second range. In particular, we reveal the influence of ion species and concentrations on the assembly. By tuning the flow rates and thus concentration profiles, we find a minimal critical salt concentration for the initiation of the assembly. Furthermore, our analysis of the surface sensitive Porod regime in the x-ray data reveals that the formation of first assembly intermediates, so-called unit length filaments, is not a one-step reaction but consists of distinct consecutive lateral association steps followed by radial compaction as well as smoothening of the surface of the full-width filament.
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http://dx.doi.org/10.1063/1.4943916DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4798992PMC
March 2016

ISPyB for BioSAXS, the gateway to user autonomy in solution scattering experiments.

Acta Crystallogr D Biol Crystallogr 2015 Jan 1;71(Pt 1):76-85. Epub 2015 Jan 1.

European Molecular Biology Laboratory, Grenoble Outstation, 71 avenue des Martyrs, CS 90181, 38042 Grenoble, France.

Logging experiments with the laboratory-information management system ISPyB (Information System for Protein crystallography Beamlines) enhances the automation of small-angle X-ray scattering of biological macromolecules in solution (BioSAXS) experiments. The ISPyB interface provides immediate user-oriented online feedback and enables data cross-checking and downstream analysis. To optimize data quality and completeness, ISPyBB (ISPyB for BioSAXS) makes it simple for users to compare the results from new measurements with previous acquisitions from the same day or earlier experiments in order to maximize the ability to collect all data required in a single synchrotron visit. The graphical user interface (GUI) of ISPyBB has been designed to guide users in the preparation of an experiment. The input of sample information and the ability to outline the experimental aims in advance provides feedback on the number of measurements required, calculation of expected sample volumes and time needed to collect the data: all of this information aids the users to better prepare for their trip to the synchrotron. A prototype version of the ISPyBB database is now available at the European Synchrotron Radiation Facility (ESRF) beamline BM29 and is already greatly appreciated by academic users and industrial clients. It will soon be available at the PETRA III beamline P12 and the Diamond Light Source beamlines I22 and B21.
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http://dx.doi.org/10.1107/S1399004714019609DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4304688PMC
January 2015

Impact of ion valency on the assembly of vimentin studied by quantitative small angle X-ray scattering.

Soft Matter 2014 Mar;10(12):2059-68

The assembly kinetics of intermediate filament (IF) proteins from tetrameric complexes to single filaments and networks depends on the protein concentration, temperature and the ionic composition of their environment. We systematically investigate how changes in the concentration of monovalent potassium and divalent magnesium ions affect the internal organization of the resulting filaments. Small angle X-ray scattering (SAXS) is very sensitive to changes in the filament cross-section such as diameter or compactness. Our measurements reveal that filaments formed in the presence of magnesium chloride differ distinctly from filaments formed in the presence of potassium chloride. The principle multi-step assembly mechanism from tetramers via unit-length filaments (ULF) to elongated filaments is not changed by the valency of ions. However, the observed differences indicate that the magnesium ions free the head domains of tetramers from unproductive interactions to allow assembly but at the same time mediate strong inter-tetrameric interactions that impede longitudinal annealing of unit-length filaments considerably, thus slowing down filament growth.
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http://dx.doi.org/10.1039/c3sm52532eDOI Listing
March 2014
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