19,179 research outputs found
DNA nano-mechanics: how proteins deform the double helix
It is a standard exercise in mechanical engineering to infer the external
forces and torques on a body from its static shape and known elastic
properties. Here we apply this kind of analysis to distorted double-helical DNA
in complexes with proteins. We extract the local mean forces and torques acting
on each base-pair of bound DNA from high-resolution complex structures. Our
method relies on known elastic potentials and a careful choice of coordinates
of the well-established rigid base-pair model of DNA. The results are robust
with respect to parameter and conformation uncertainty. They reveal the complex
nano-mechanical patterns of interaction between proteins and DNA. Being
non-trivially and non-locally related to observed DNA conformations, base-pair
forces and torques provide a new view on DNA-protein binding that complements
structural analysis.Comment: accepted for publication in JCP; some minor changes in response to
review 18 pages, 5 figure + supplement: 4 pages, 3 figure
Imaging and 3D reconstruction of membrane protein complexes by cryo-electron microscopy and single particle analysis
Cryo-electron microscopy (cryo-EM) in combination with single particle image processing and volume reconstruction is a powerful technology to obtain medium-resolution structures of large protein complexes, which are extremely diļ¬cult to crystallize and not amenable to NMR studies due to size limitation. Depending on the stability and stiļ¬ness as well as on the symmetry of the complex, three-dimensional reconstructions at a resolution of 10-30 Ė can be achieved. In this range of resolution, we may not be able to answer A chemical questions at the level of atomic interactions, but we can gain detailed insight into the macromolecular architecture of large multi-subunit complexes and their mechanisms of action. In this thesis, several prevalently large membrane protein complexes of great physiological importance were examined by various electron microscopy techniques and single particle image analysis. The core part of my work consists in the imaging of a mammalian V-ATPase, frozen-hydrated in amorphous ice and of the completion of the ļ¬rst volume reconstruction of this type of enzyme, derived from cryo-EM images. This ubiquitous rotary motor is essential in every eukaryotic cell and is of high medical importance due to its implication in various diseases such as osteoporosis, skeletal cancer and kidney disorders. My contribution to the second and third paper concerns the volume reconstruction of two bacterial outer membrane pore complexes from cryo-EM images recorded by my colleague Mohamed Chami. PulD from Klebsiella oxytoca constitutes a massive translocating pore capable of transporting a fully folded cell surface protein PulA through the membrane. It is part of the Type II secretion system, which is common for Gram-negative bacteria. The second volume regards ClyA, a pore-forming heamolytic toxin of virulent Escherichia coli and Salmonella enterica strains that kill target cells by inserting pores into their membranes. To the last two papers, I contributed with cryo-negative stain imaging of the cell division protein DivIVA from Bacillus subtilis and with image processing of the micrographs displaying the siderophore receptor FrpB from Neisseria meningitidis
New Light on Molecular and Materials Complexity: 4D Electron Imaging
In this Perspective, 4D electron imaging is highlighted, after introducing some concepts, with an overview of selected applications that span chemical reactions, molecular interfaces, phase transitions, and nano(micro)mechanical systems. With the added dimension of time in microscopy, diffraction, and electron-energy-loss spectroscopy, the focus is on direct visualization of structural dynamics with atomic and nanoscale resolution in the four dimensions of space and time. This contribution provides an expose of emerging developments and an outlook on future applications in materials and biological sciences
- ā¦