scholarly journals A hold-and-feed mechanism drives directional DNA loop extrusion by condensin

2021 ◽  
Author(s):  
Indra A Shaltiel ◽  
Sumanjit Datta ◽  
Léa Lecomte ◽  
Markus Hassler ◽  
Marc Kschonsak ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Single Molecule ◽  
Protein Complexes ◽  
Double Helix ◽  
Power Stroke ◽  
Reaction Cycle ◽  
Cryo Electron Microscopy ◽  
Condensin Complex ◽  
Dna Loop ◽  
Dna Double Helix

SMC protein complexes structure genomes by extruding DNA loops, but the molecular mechanism that underlies their activity has remained unknown. We show that the active condensin complex entraps the bases of a DNA loop in two separate chambers. Single-molecule and cryo-electron microscopy provide evidence for a power-stroke movement at the first chamber that feeds DNA into the SMC-kleisin ring upon ATP binding, while the second chamber holds on upstream of the same DNA double helix. Unlocking the strict separation of 'motor' and 'anchor' chambers turns condensin from a one-sided into a bidirectional DNA loop extruder. We conclude that the orientation of two topologically bound DNA segments during the course of the SMC reaction cycle determines the directionality of DNA loop extrusion.

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 Real-time imaging of DNA loop extrusion by condensin

Science ◽  
2018 ◽  
Vol 360 (6384) ◽  
pp. 102-105 ◽  
Author(s):  
Mahipal Ganji ◽  
Indra A. Shaltiel ◽  
Shveta Bisht ◽  
Eugene Kim ◽  
Ana Kalichava ◽  
...  
Keyword(s):  
Real Time ◽  
Adenosine Triphosphate ◽  
Genome Organization ◽  
Protein Complexes ◽  
Dna Loops ◽  
Base Pairs ◽  
Real Time Imaging ◽  
Condensin Complex ◽  
Dna Loop

It has been hypothesized that SMC protein complexes such as condensin and cohesin spatially organize chromosomes by extruding DNA into large loops. We directly visualized the formation and processive extension of DNA loops by yeast condensin in real time. Our findings constitute unambiguous evidence for loop extrusion. We observed that a single condensin complex is able to extrude tens of kilobase pairs of DNA at a force-dependent speed of up to 1500 base pairs per second, using the energy of adenosine triphosphate hydrolysis. Condensin-induced loop extrusion was strictly asymmetric, which demonstrates that condensin anchors onto DNA and reels it in from only one side. Active DNA loop extrusion by SMC complexes may provide the universal unifying principle for genome organization.

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.

scholarly journals Cryo-electron microscopy and single molecule fluorescent microscopy detect CD4 receptor induced HIV size expansion prior to cell entry

Virology ◽  
2015 ◽  
Vol 486 ◽  
pp. 121-133 ◽  
Author(s):  
Son Pham ◽  
Thibault Tabarin ◽  
Megan Garvey ◽  
Corinna Pade ◽  
Jérémie Rossy ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Single Molecule ◽  
Fluorescent Microscopy ◽  
Cell Entry ◽  
Cd4 Receptor ◽  
Cryo Electron Microscopy

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].

Direct Single-Molecule Observations of Local Denaturation of a DNA Double Helix under a Negative Supercoil State

2015 ◽  
Vol 87 (6) ◽  
pp. 3490-3497 ◽  
Author(s):  
Shunsuke Takahashi ◽  
Shinya Motooka ◽  
Tomohiro Usui ◽  
Shohei Kawasaki ◽  
Hidefumi Miyata ◽  
...  
Keyword(s):  
Single Molecule ◽  
Double Helix ◽  
Dna Double Helix

scholarly journals Mechanism of RPA-Facilitated Processive DNA Unwinding by the Eukaryotic CMG Helicase

2019 ◽  
Author(s):  
Hazal B. Kose ◽  
Sherry Xie ◽  
George Cameron ◽  
Melania S. Strycharska ◽  
Hasan Yardimci
Keyword(s):  
Single Molecule ◽  
Replication Fork ◽  
Double Helix ◽  
Dna Unwinding ◽  
Helicase Activity ◽  
Replicative Helicase ◽  
Double Stranded Dna ◽  
Order Of Magnitude ◽  
Dna Strand ◽  
Dna Double Helix

AbstractThe DNA double helix is unwound by the Cdc45/Mcm2-7/GINS (CMG) complex at the eukaryotic replication fork. While isolated CMG unwinds duplex DNA very slowly, its fork unwinding rate is stimulated by an order of magnitude by single-stranded DNA binding protein, RPA. However, the molecular mechanism by which RPA enhances CMG helicase activity remained elusive. Here, we demonstrate that engagement of CMG with parental double-stranded DNA (dsDNA) at the replication fork impairs its helicase activity, explaining the slow DNA unwinding by isolated CMG. Using single-molecule and ensemble biochemistry, we show that binding of RPA to the excluded DNA strand prevents duplex engagement by the helicase and speeds up CMG-mediated DNA unwinding. When stalled due to dsDNA interaction, DNA rezipping-induced helicase backtracking re-establishes productive helicase-fork engagement underscoring the significance of plasticity in helicase action. Together, our results elucidate the dynamics of CMG at the replication fork and reveal how other replisome components can mediate proper DNA engagement by the replicative helicase to achieve efficient fork progression.

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