Tag: Cryo-Electron Microscopy

Freezing a Moment in Time: Snapshots of Cryo-EM Research

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To get a look at cell components that are too small to see with a normal light microscope, scientists often use cryo-electron microscopy (cryo-EM). As the prefix cryo- means “cold” or “freezing,” cryo-EM involves rapidly freezing a cell, virus, molecular complex, or other structure to prevent water molecules from forming crystals. This preserves the sample in its natural state and keeps it still so that it can be imaged with an electron microscope, which uses beams of electrons instead of light. Some electrons are scattered by the sample, while others pass through it and through magnetic lenses to land on a detector and form an image.

Typically, samples contain many copies of the object a scientist wants to study, frozen in a range of orientations. Researchers take images of these various positions and combine them into a detailed 3D model of the structure. Electron microscopes allow us to see much smaller structures than light microscopes do because the wavelengths of electrons are much shorter than the wavelength of light. NIGMS-funded researchers are using cryo-EM to investigate a range of scientific questions.

Caught in Translation

One cluster that is yellow, purple, and orange and another that is beige, purple, and green. 3D reconstructions of two stages in the assembly of the bacterial ribosome created from time-resolved cryo-EM images. Credit: Joachim Frank, Columbia University.

Joachim Frank, Ph.D., a professor of biochemistry and molecular biophysics and of biological sciences at Columbia University in New York, New York, along with two other researchers, won the 2017 Nobel Prize in Chemistry for developing cryo.

Dr. Frank’s lab focuses on the process of translation, where structures called ribosomes turn genetic instructions into proteins, which are needed for many chemical reactions that support life. Recently, Dr. Frank has adopted and further developed a technique called time-resolved cryo-EM. This method captures images of short-lived states in translation that disappear too quickly (after less than a second) for standard cryo-EM to capture. The ability to fully visualize translation could help researchers identify errors in the process that lead to disease and also to develop treatments.

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Cool Images: The Hidden Beauty Inside Plants

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Spring brings with it a wide array of beautiful flowers, but the interior structures of plants can be just as stunning. Using powerful microscopes, researchers can peek into the many molecular bits and pieces that make up plants. Check out these cool plant images from our Image and Video Gallery that NIGMS-funded scientists created while doing their research.

Several round structures that are yellow at the center and pink and purple around the edges and have honeycomb-like interiors. Credit: Arun Sampathkumar and Elliot Meyerowitz, California Institute of Technology.

In plants and animals, stem cells can transform into a variety of different cell types. The stem cells at the growing tip of this Arabidopsis plant will soon become flowers. Cellular and molecular biologists frequently study Arabidopsis because it grows rapidly (its entire life cycle is only 6 weeks), produces lots of seeds, and has a genome that’s easy to manipulate.

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Revealing a Piece of Cilia’s Puzzle

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A multicolored tube made up of small dots with three sets of appendages attached along its length. A partial model of a doublet microtubule. Credit: Veronica Falconieri.

Cilia (cilium in singular) are complex organelles found on all of our cells except red blood cells. Their rhythmic beating moves fluid or materials over the cell to help transport food and oxygen or remove debris. For example, cilia in our windpipe prevent bacteria and mucous from traveling to the lungs. Some pick up signals like antennae, such as cilia in our ears that help detect sounds. One component of cilia is the doublet microtubule, a major part of cilia’s skeleton that gives it strength and rigidity.

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Researchers Score Goal with New Atomic-Scale Model of Salmonella-Infecting Virus

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An atomic-scale model of a virus that infects the Salmonella bacterium. Credit: C. Hryc and the Chiu Lab, Baylor College of Medicine.

This sphere could be a prototype design for the 2018 World Cup official match soccer ball, but you won’t see it dribbled around any soccer fields. The image is actually an atomic-scale model of a virus that infects the Salmonella bacterium. Like a soccer ball, both are approximately spherical shapes created by a combination of hexagonal (six-sided) and pentagonal (five-sided) units. Wah Chiu, a biochemist at Baylor College of Medicine, and his colleagues used new computational methods to construct the model from more than 20,000 cryo-electron microscopy (cryo-EM) images. Cryo-EM is a sophisticated technique that uses electron beams for visualizing frozen samples of proteins and other biological specimens.

The researchers’ model, published in a recent issue of PNAS, shows the virus’ protein shell, or capsid, that encloses the virus’ genetic material. Each color shows capsid proteins having the same interactions with their neighbors. The fine resolution allowed researchers to identify the protein interactions essential to building a stable shell. They developed a new approach to checking the accuracy and reliability of the virus model and reporting what parts are the most certain. The approach could be used to evaluate other complex biological structures, potentially leading to better quality models and new avenues for drug design and development.

This research is funded in part by NIH under grants R01GM079429, P01GM063210, P41GM103832, PN2EY016525, T15LM007093.

Cool Tools: Pushing the Limits of High-Resolution Microscopy

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Cell biologists would love to shrink themselves down and actually see, touch and hear the inner workings of cells. Because that’s impossible, they have developed an ever-growing collection of microscopes to study cellular innards from the outside. Using these powerful tools, researchers can exhaustively inventory the molecular bits and pieces that make up cells, eavesdrop on cellular communication and spy on cells as they adapt to changing environments.

In recent years, scientists have developed new cellular imaging techniques that allow them to visualize samples in ways and at levels of detail never before possible. Many of these techniques build upon the power of electron microscopy (EM) to see ever smaller details.

Unlike traditional light microscopy, EM uses electrons, not light, to create an image. To do so, EM accelerates electrons in a vacuum, shoots them out of an electron gun and focuses them with doughnut-shaped magnets onto a sample. When electrons bombard the sample, some pass though without being absorbed while others are scattered. The transmitted electrons land on a detector and produce an image, just as light strikes a detector (or film) in a camera to create a photograph.

This image, showing a single protein molecule, is a montage. It was created to demonstrate how dramatically cryo-EM has improved in recent years. In the past, cryo-EM was only able to obtain a blobby approximation of a molecule’s shape, like that shown on the far left. Now, the technique yields exquisitely detailed images in which individual atoms are nearly visible (far right). Color is artificially applied. Credit: Veronica Falconieri, Subramaniam Lab, National Cancer Institute.

Transmission electron microscopes can magnify objects more than 10 million times, enabling scientists to see the outline and some details of cells, viruses and even some large molecules. A relatively new form of transmission electron microscopy called cryo-EM enables scientists to view specimens in their natural or near-natural state without the need for dyes or stains.

In cryo-EM—the prefix cry- means “cold” or “freezing”—scientists freeze a biological sample so rapidly that water molecules do not have time to form ice crystals, which could shove cellular materials out of their normal place. Cold samples are more stable and can be imaged many times over, allowing researchers to iteratively refine the image, remove artifacts and produce even sharper images than ever before. Continue reading “Cool Tools: Pushing the Limits of High-Resolution Microscopy”

Viral Views: New Insights on Infection Strategies

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The following images show a few ways in which cutting-edge research tools are giving us clearer views of viruses—and possible ways to disarm them. The examples, which highlight work involving HIV and the coronavirus, were funded in part by our Biomedical Technology Research Resources program.

Uncloaking HIV’s Camouflage

HIV capsid with (right, red) and without (left) a camouflaging human protein.
HIV capsid with (right, red) and without (left) a camouflaging human protein. Credit: Juan R. Perilla, Klaus Schulten and the Theoretical and Computational Biophysics Group, University of Illinois at Urbana-Champaign.

To sneak past our immune defenses and infect human cells, HIV uses a time-honored strategy—disguise. The virus’ genome is enclosed in a protein shell called a capsid (on left) that’s easily recognized and destroyed by the human immune system. To evade this fate, the chrysalis-shaped capsid cloaks itself with a human protein known as cyclophilin A (in red, on right). Camouflaged as human, the virus gains safe passage into and through a human cell to deposit its genetic material in the nucleus and start taking control of cellular machinery.

Biomedical and technical experts teamed up to generate these HIV models at near-atomic resolution. First, structural biologists at the Pittsburgh Center for HIV Protein Interactions Exit icon used a technique called cryo-electron microscopy (cryo-EM) to get information on the shape of an HIV capsid as well as the capsid-forming proteins’ connections to each other and to cyclophilin A. Then experts at the Resource for Macromolecular Modeling and Bioinformatics fed the cryo-EM data into their visualization and simulation programs to computationally model the physical interactions among every single atom of the capsid and the cyclophilin A protein. The work revealed a previously unknown site where cyclophilin A binds to the capsid, offering new insights on the biology of HIV infection. Continue reading “Viral Views: New Insights on Infection Strategies”

Cryo-Electron Microscopy Reveals Molecules in Ever Greater Detail

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The molecular visualization technique known as cryo-electron microscopy (cryo-EM) was recently named the “2015 Method of the Year” by the journal Nature Methods. In a recent blog post, NIH Director Francis Collins explains how the technology works and just how rapidly it has advanced. He writes, “Today’s cryo-EM is so powerful that researchers can almost make out individual atoms!” He also notes, “These dramatic advances serve as a reminder of the ways in which basic technological innovation can open new realms of scientific possibility.”

We fund many scientists who are developing and applying cryo-EM to bring the details of biological structures into unprecedented focus. Here are two examples of their work and its potential impact. Continue reading “Cryo-Electron Microscopy Reveals Molecules in Ever Greater Detail”