Gctf: Real-time CTF determination and correction

Journal of Structural Biology, Год журнала: 2015, Номер 193(1), С. 1 - 12

Опубликована: Ноя. 20, 2015

Accurate estimation of the contrast transfer function (CTF) is critical for a near-atomic resolution cryo electron microscopy (cryoEM) reconstruction. Here, GPU-accelerated computer program, Gctf, accurate and robust, real-time CTF determination presented. The main target Gctf to maximize cross-correlation simulated with logarithmic amplitude spectra (LAS) observed micrographs after background subtraction. Novel approaches in improve both speed accuracy. In addition GPU acceleration (e.g. 10–50×), fast ‘1-dimensional search plus 2-dimensional refinement (1S2R)’ procedure further speeds up Gctf. Based on global determination, local defocus each particle single frames movies accurately refined, which improves parameters all particles subsequent image processing. diagnosis method using equiphase averaging (EPA) self-consistency verification procedures have also been implemented program practical use, especially aims an independent outputs can be easily imported into other cryoEM software such as Relion (Scheres, 2012) Frealign (Grigorieff, 2007). results from several representative datasets are shown discussed this paper.

Язык: Английский

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DNA Origami: Scaffolds for Creating Higher Order Structures

Chemical Reviews, Год журнала: 2017, Номер 117(20), С. 12584 - 12640

Опубликована: Июнь 12, 2017

DNA has become one of the most extensively used molecular building blocks for engineering self-assembling materials. origami is a technique that uses hundreds short oligonucleotides, called staple strands, to fold long single-stranded DNA, which scaffold strand, into various designer nanoscale architectures. dramatically improved complexity and scalability nanostructures. Due its high degree customization spatial addressability, provides versatile platform with engineer structures devices can sense, compute, actuate. These capabilities open up opportunities broad range applications in chemistry, biology, physics, material science, computer science have often required programmed control molecules atoms three-dimensional (3D) space. This review comprehensive survey recent developments structure, design, assembly, directed self-assembly, as well applications.

Язык: Английский

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Half a century of amyloids: past, present and future

Chemical Society Reviews, Год журнала: 2020, Номер 49(15), С. 5473 - 5509

Опубликована: Янв. 1, 2020

Amyloid diseases are global epidemics with profound health, social and economic implications yet remain without a cure. This dire situation calls for research into the origin pathological manifestations of amyloidosis to stimulate continued development new therapeutics. In basic science engineering, cross-β architecture has been constant thread underlying structural characteristics functional amyloids, realizing that amyloid structures can be both in nature fuelled innovations artificial whose use today ranges from water purification 3D printing. At conclusion half century since Eanes Glenner's seminal study amyloids humans, this review commemorates occasion by documenting major milestones date, perspectives biology, biophysics, medicine, microbiology, engineering nanotechnology. We also discuss challenges opportunities drive interdisciplinary field moving forward.

Язык: Английский

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Protein Ensembles: How Does Nature Harness Thermodynamic Fluctuations for Life? The Diverse Functional Roles of Conformational Ensembles in the Cell

Chemical Reviews, Год журнала: 2016, Номер 116(11), С. 6516 - 6551

Опубликована: Янв. 25, 2016

All soluble proteins populate conformational ensembles that together constitute the native state. Their fluctuations in water are intrinsic thermodynamic phenomena, and distributions of states on energy landscape determined by statistical thermodynamics; however, they optimized to perform their biological functions. In this review we briefly describe advances free studies protein ensembles. Experimental (nuclear magnetic resonance, small-angle X-ray scattering, single-molecule spectroscopy, cryo-electron microscopy) computational (replica-exchange molecular dynamics, metadynamics, Markov state models) approaches have made great progress recent years. These address challenging characterization highly flexible heterogeneous We focus structural aspects distributions, from collective motions single- multi-domain proteins, intrinsically disordered multiprotein complexes. Importantly, highlight illustrate functional adjustment crowded cellular environment. center role ensemble recognition small- macro-molecules (protein RNA/DNA) emphasize emerging concepts dynamics enzyme catalysis. Overall, link fundamental physicochemical principles behavior network its regulation.

Язык: Английский

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EMDataBank unified data resource for 3DEM

Nucleic Acids Research, Год журнала: 2015, Номер 44(D1), С. D396 - D403

Опубликована: Ноя. 17, 2015

Three-dimensional Electron Microscopy (3DEM) has become a key experimental method in structural biology for broad spectrum of biological specimens from molecules to cells. The EMDataBank project provides unified portal deposition, retrieval and analysis 3DEM density maps, atomic models associated metadata (emdatabank.org). We provide here an overview the rapidly growing data archives, which include maps EM Data Bank map-derived Protein Bank. In addition, we describe progress approaches toward development validation protocols methods, working with scientific community, order create pipeline data.

Язык: Английский

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Cryo-EM structure of haemoglobin at 3.2 Å determined with the Volta phase plate

Nature Communications, Год журнала: 2017, Номер 8(1)

Опубликована: Июнь 30, 2017

With the advent of direct electron detectors, perspectives cryo-electron microscopy (cryo-EM) have changed in a profound way. These cameras are superior to previous detectors coping with intrinsically low contrast and beam-induced motion radiation-sensitive organic materials embedded amorphous ice, hence they enabled structure determination many macromolecular assemblies atomic or near-atomic resolution. Nevertheless, there still limitations one them is size target structure. Here, we report use Volta phase plate determining human haemoglobin (64 kDa) at 3.2 Å. Our results demonstrate that this method can be applied complexes significantly smaller than those previously studied by conventional defocus-based approaches. Cryo-EM now close becoming fast cost-effective alternative crystallography for high-resolution protein determination.

Язык: Английский

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Conformational Activation Promotes CRISPR-Cas12a Catalysis and Resetting of the Endonuclease Activity

Cell, Год журнала: 2018, Номер 175(7), С. 1856 - 1871.e21

Опубликована: Ноя. 29, 2018

Язык: Английский

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Cryo-Electron Microscopy Methodology: Current Aspects and Future Directions

Trends in Biochemical Sciences, Год журнала: 2019, Номер 44(10), С. 837 - 848

Опубликована: Май 8, 2019

Язык: Английский

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High-resolution structure determination of sub-100 kDa complexes using conventional cryo-EM

Nature Communications, Год журнала: 2019, Номер 10(1)

Опубликована: Март 4, 2019

Determining high-resolution structures of biological macromolecules amassing less than 100 kilodaltons (kDa) has been a longstanding goal the cryo-electron microscopy (cryo-EM) community. While Volta phase plate enabled visualization specimens in this size range, instrumentation is not yet fully automated and can present technical challenges. Here, we show that conventional defocus-based cryo-EM methodologies be used to determine kDa using transmission electron microscope operating at 200 keV coupled with direct detector. Our ~2.7 Å structure alcohol dehydrogenase (82 kDa) proves bound ligands resolved high fidelity enable investigation drug-target interactions. ~2.8 ~3.2 methemoglobin demonstrate distinct conformational states identified within dataset for proteins as small 64 kDa. Furthermore, provide sub-nanometer sub-50 protein.

Язык: Английский

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The myosin mesa and the basis of hypercontractility caused by hypertrophic cardiomyopathy mutations

Nature Structural & Molecular Biology, Год журнала: 2017, Номер 24(6), С. 525 - 533

Опубликована: Май 8, 2017

Язык: Английский

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