scholarly journals Cryo-electron microscopy structure of a nucleosome-bound SWI/SNF chromatin remodeling complex

2019 ◽  
Author(s):  
Yan Han ◽  
Alexis A Reyes ◽  
Sara Malik ◽  
Yuan He
Keyword(s):  
Electron Microscopy ◽  
Chromatin Remodeling ◽  
Atp Hydrolysis ◽  
Protein Complexes ◽  
Gene Activation ◽  
The Body ◽  
Cellular Processes ◽  
Cryo Electron Microscopy ◽  
Chromatin Remodeling Complex ◽  
High Resolution Structure

AbstractThe multi-subunit chromatin remodeling complex SWI/SNF1–3 is highly conserved from yeast to humans and plays critical roles in various cellular processes including transcription and DNA damage repair4, 5. It uses the energy from ATP hydrolysis to remodel chromatin structure by sliding and evicting the histone octamer6–10, creating DNA regions that become accessible to other essential protein complexes. However, our mechanistic understanding of the chromatin remodeling activity is largely hindered by the lack of a high-resolution structure of any complex from this family. Here we report the first structure of SWI/SNF from the yeast S. cerevisiae bound to a nucleosome at near atomic resolution determined by cryo-electron microscopy (cryo-EM). In the structure, the Arp module is sandwiched between the ATPase and the Body module of the complex, with the Snf2 HSA domain connecting all modules. The HSA domain also extends into the Body and anchors at the opposite side of the complex. The Body contains an assembly scaffold composed of conserved subunits Snf12 (SMARCD/BAF60), Snf5 (SMARCB1/BAF47/ INI1) and an asymmetric dimer of Swi3 (SMARCC/BAF155/170). Another conserved subunit Swi1 (ARID1/BAF250) folds into an Armadillo (ARM) repeat domain that resides in the core of the SWI/SNF Body, acting as a molecular hub. In addition to the interaction between Snf2 and the nucleosome, we also observed interactions between the conserved Snf5 subunit and the histones at the acidic patch, which could serve as an anchor point during active DNA translocation. Our structure allows us to map and rationalize a subset of cancer-related mutations in the human SWI/SNF complex and propose a model of how SWI/SNF recognizes and remodels the +1 nucleosome to generate nucleosome-depleted regions during gene activation11–13.

scholarly journals PRISM-EM: template interface-based modelling of multi-protein complexes guided by cryo-electron microscopy density maps

2016 ◽  
Vol 72 (10) ◽  
pp. 1137-1148 ◽  
Author(s):  
Guray Kuzu ◽  
Ozlem Keskin ◽  
Ruth Nussinov ◽  
Attila Gursoy
Keyword(s):  
Electron Microscopy ◽  
Protein Complexes ◽  
Three Dimensional ◽  
Protein Assemblies ◽  
New Approach ◽  
Cellular Processes ◽  
Cryo Electron Microscopy ◽  
Density Maps ◽  
Crystallographic Structures ◽  
Hiv 1

The structures of protein assemblies are important for elucidating cellular processes at the molecular level. Three-dimensional electron microscopy (3DEM) is a powerful method to identify the structures of assemblies, especially those that are challenging to study by crystallography. Here, a new approach, PRISM-EM, is reported to computationally generate plausible structural models using a procedure that combines crystallographic structures and density maps obtained from 3DEM. The predictions are validated against seven available structurally different crystallographic complexes. The models display mean deviations in the backbone of <5 Å. PRISM-EM was further tested on different benchmark sets; the accuracy was evaluated with respect to the structure of the complex, and the correlation with EM density maps and interface predictions were evaluated and compared with those obtained using other methods. PRISM-EM was then used to predict the structure of the ternary complex of the HIV-1 envelope glycoprotein trimer, the ligand CD4 and the neutralizing protein m36.

scholarly journals Von der „Blobology“ zur atomaren Auflösung der Kryo-Elektronenmikroskopie

BIOspektrum ◽  
2020 ◽  
Vol 26 (7) ◽  
pp. 710-713
Author(s):  
Holger Stark
Keyword(s):  
Electron Microscopy ◽  
High Resolution ◽  
Structure Determination ◽  
Protein Complexes ◽  
Pharmaceutical Research ◽  
New Drugs ◽  
Cryo Electron Microscopy ◽  
High Resolution Structure ◽  
First Time

AbstractIt took almost a century to develop electron microscopy into a powerful method for high-resolution structure determination of proteins. Technical improvements in microscopy, detector technology, and image processing software contributed to the exponential growth of high-resolution structures of protein complexes determined by cryo-electron microscopy in recent years. We now succeeded in breaking another resolution barrier in cryo-electron microscopy and for the first time in achieving true atomic resolution, where single atoms in the protein can indeed be visualized individually. These improvements in cryo-EM indicate that the method will continue to gain importance, not only as a method for structure determination but also in the development of new drugs in pharmaceutical research.

scholarly journals Substrate-engaged 26S proteasome structures reveal mechanisms for ATP-hydrolysis–driven translocation

Science ◽  
2018 ◽  
Vol 362 (6418) ◽  
pp. eaav0725 ◽  
Author(s):  
Andres H. de la Peña ◽  
Ellen A. Goodall ◽  
Stephanie N. Gates ◽  
Gabriel C. Lander ◽  
Andreas Martin
Keyword(s):  
Electron Microscopy ◽  
Conformational Changes ◽  
Adenosine Triphosphatase ◽  
Atp Hydrolysis ◽  
26S Proteasome ◽  
Phosphate Release ◽  
Protein Substrates ◽  
Cellular Processes ◽  
Cryo Electron Microscopy ◽  
Aaa Atpases

The 26S proteasome is the primary eukaryotic degradation machine and thus is critically involved in numerous cellular processes. The heterohexameric adenosine triphosphatase (ATPase) motor of the proteasome unfolds and translocates targeted protein substrates into the open gate of a proteolytic core while a proteasomal deubiquitinase concomitantly removes substrate-attached ubiquitin chains. However, the mechanisms by which ATP hydrolysis drives the conformational changes responsible for these processes have remained elusive. Here we present the cryo–electron microscopy structures of four distinct conformational states of the actively ATP-hydrolyzing, substrate-engaged 26S proteasome. These structures reveal how mechanical substrate translocation accelerates deubiquitination and how ATP-binding, -hydrolysis, and phosphate-release events are coordinated within the AAA+ (ATPases associated with diverse cellular activities) motor to induce conformational changes and propel the substrate through the central pore.

CRYO-Electron Microscopy of the Acto-Myosin-ATP Complex

1990 ◽  
Vol 48 (1) ◽  
pp. 248-249
Author(s):  
John Trinickt ◽  
Howard White
Keyword(s):  
Electron Microscopy ◽  
Atp Hydrolysis ◽  
Myosin Head ◽  
Bound State ◽  
Cross Linking ◽  
Weakly Bound ◽  
Cryo Electron Microscopy ◽  
Myosin Subfragment 1 ◽  
Attached State ◽  
High Fraction

The primary force of muscle contraction is thought to involve a change in the myosin head whilst attached to actin, the energy coming from ATP hydrolysis. This change in attached state could either be a conformational change in the head or an alteration in the binding angle made with actin. A considerable amount is known about one bound state, the so-called strongly attached state, which occurs in the presence of ADP or in the absence of nucleotide. In this state, which probably corresponds to the last attached state of the force-producing cycle, the angle between the long axis myosin head and the actin filament is roughly 45°. Details of other attached states before and during power production have been difficult to obtain because, even at very high protein concentration, the complex is almost completely dissociated by ATP. Electron micrographs of the complex in the presence of ATP have therefore been obtained only after chemically cross-linking myosin subfragment-1 (S1) to actin filaments to prevent dissociation. But it is unclear then whether the variability in attachment angle observed is due merely to the cross-link acting as a hinge.We have recently found low ionic-strength conditions under which, without resorting to cross-linking, a high fraction of S1 is bound to actin during steady state ATP hydrolysis. The structure of this complex is being studied by cryo-electron microscopy of hydrated specimens. Most advantages of frozen specimens over ambient temperature methods such as negative staining have already been documented. These include improved preservation and fixation rates and the ability to observe protein directly rather than a surrounding stain envelope. In the present experiments, hydrated specimens have the additional benefit that it is feasible to use protein concentrations roughly two orders of magnitude higher than in conventional specimens, thereby reducing dissociation of weakly bound complexes.

scholarly journals Application of Cryo-Electron Microscopy on Drug Discovery

2021 ◽  
Vol 27 (S1) ◽  
pp. 3250-3250
Author(s):  
Viswanath Vittaladevaram ◽  
Kranthi Kuruti
Keyword(s):  
Electron Microscopy ◽  
Protein Complexes ◽  
Lead Discovery ◽  
Biologically Relevant ◽  
Biological Targets ◽  
3 Dimensional ◽  
Drug Molecules ◽  
Cryo Electron Microscopy ◽  
Therapeutic Molecules ◽  
Novel Drug

AbstractThe key aspect for development of novel drug molecules is to perform structural determination of target molecule associated with its ligand. One such tool that provides insights towards structure of molecule is Cryo-electron microscopy which covers biological targets that are intractable. Examination of proteins can be carried out in native state, as the samples are frozen at -175 degree Celsius i.e. cryogenic temperatures. In addition to this, there were no limits for molecular and functional structures of proteins that can be imagined in 3-dimensional form. This includes ligands which unravel mechanisms that are biologically relevant. This will enable to better understand the mechanisms that are used for development of new therapeutics. Application of Cryo-electron microscopy is not limited to protein complexes and is considered as non-specific. Intervention of Cryo-EM would allow to analyse the structures and also able to dissect the interaction with therapeutic molecules. The study determines the usage of cryo-EM for providing resolutions that are acceptable for lead discovery. It also provides support for lead optimization and also for discovery of vaccines and therapeutics.

scholarly journals Rapid high-resolution structure analysis of small, biotechnologically relevant enzymes by cryo-electron microscopy

2021 ◽  
Author(s):  
Nicole Dimos ◽  
Carl P.O. Helmer ◽  
Andrea M. Chanique ◽  
Markus C. Wahl ◽  
Robert Kourist ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
High Resolution ◽  
Structural Biology ◽  
Structure Analysis ◽  
Cryo Electron Microscopy ◽  
High Resolution Structure ◽  
Fast Development ◽  
Pharmaceutical Industries ◽  
Exquisite Specificity ◽  
Resolution Structure

Enzyme catalysis has emerged as a key technology for developing efficient, sustainable processes in the chemical, biotechnological and pharmaceutical industries. Plants provide large and diverse pools of biosynthetic enzymes that facilitate complex reactions, such as the formation of intricate terpene carbon skeletons, with exquisite specificity. High-resolution structural analysis of these enzymes is crucial to understand their mechanisms and modulate their properties by targeted engineering. Although cryo-electron microscopy (cryo-EM) has revolutionized structural biology, its applicability to high-resolution structure analysis of comparatively small enzymes is so far largely unexplored. Here, we show that cryo-EM can reveal the structures of ~120 kDa plant borneol dehydrogenases at or below 2 Å resolution, paving the way for the fast development of new biocatalysts that provide access to bioactive terpenes and terpenoids.

scholarly journals Opportunities and challenges for assigning cofactors in cryo-EM density maps of chlorophyll-containing proteins

2020 ◽  
Vol 3 (1) ◽  
Author(s):  
Christopher J. Gisriel ◽  
Jimin Wang ◽  
Gary W. Brudvig ◽  
Donald A. Bryant
Keyword(s):  
Electron Microscopy ◽  
Electrostatic Potential ◽  
Protein Function ◽  
Protein Complexes ◽  
Chemical Environment ◽  
Cryo Electron Microscopy ◽  
Functional Differences ◽  
Density Maps ◽  
Structural Differences

AbstractThe accurate assignment of cofactors in cryo-electron microscopy maps is crucial in determining protein function. This is particularly true for chlorophylls (Chls), for which small structural differences lead to important functional differences. Recent cryo-electron microscopy structures of Chl-containing protein complexes exemplify the difficulties in distinguishing Chl b and Chl f from Chl a. We use these structures as examples to discuss general issues arising from local resolution differences, properties of electrostatic potential maps, and the chemical environment which must be considered to make accurate assignments. We offer suggestions for how to improve the reliability of such assignments.

Molecular dynamics of protein complexes from four-dimensional cryo-electron microscopy

2004 ◽  
Vol 147 (3) ◽  
pp. 291-301 ◽  
Author(s):  
J.Bernard Heymann ◽  
James F. Conway ◽  
Alasdair C. Steven
Keyword(s):  
Electron Microscopy ◽  
Molecular Dynamics ◽  
Protein Complexes ◽  
Cryo Electron Microscopy

scholarly journals PRISM-EM: template interface-based modelling of multi-protein complexes guided by cryo-electron microscopy density maps. Corrigendum

2018 ◽  
Vol 74 (1) ◽  
pp. 65-66
Author(s):  
Guray Kuzu ◽  
Ozlem Keskin ◽  
Ruth Nussinov ◽  
Attila Gursoy
Keyword(s):  
Electron Microscopy ◽  
Protein Complexes ◽  
Acta Cryst ◽  
Cryo Electron Microscopy ◽  
Density Maps

A revised Table 6 and Supporting Information are provided for the article by Kuzuet al.[(2016),Acta Cryst.D72, 1137–1148].

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