Cryo-Electron Microscopy (Cryo-EM) A Key Element in Recent Research
Cryo-electron microscopy, called Cryo-EM, has been around for a while. The first model using it was deposited with the PDB (Protein Data Bank) in 1997, and the number of images has grown since. The technique is a type of electron microscopy in which the sample is rapidly cooled so the water molecules don’t have time to crystallize. The resulting models are easier to visualize than in other types of EM.
Here are examples of recent uses of Cryo-EM.
Researchers at the University of Glasgow used Cryo-EM to show detailed structural images of the common herpes virus. The research was published in PLOS Biology. The herpes family of viruses cause a variety of diseases ranging from cold sores to chicken pox, as well as some cancers and other diseases. But the size of the virus’ capsid is about 1/10,000th of a millimeter in diameter, which has made it difficult for researchers to visualize it.
The scientists used Cryo-EM to create images of the herpes virus, which showed the architecture of a motor-like assembly called a portal. Herpesviruses insert their DNA through the portal using preassembled capsids, which are the shell that contains the DNA (or RNA, in RNA viruses).
“Cry-electron microscopy, combined with new computational image processing methods, allowed us to reveal the detailed structure of the unique machinery by which the virus packs DNA into the capsid,” said David Bhella, lead author of the study, in a statement. “The DNA is packed very tightly, reaching a pressure similar to that inside a bottle of champagne. We hope that this study will eventually lead to the development of new medicines to treat acute herpesvirus infections, through the design of drugs that will block the action of the portal motor.”
Ryan Hibbs, with the University of Texas Southwestern Medical Center in Dallas, published work in the June 27 issue of Nature that used Cryo-EM to image a human GABAA receptor. GABAA receptors have five subunits that work together to create a channel that lets chloride ions flow into neurons. The five subunits come from a pool of 19. The technology had been used previously to take images of a GABAA receptor built of five B3 subunits, but that are not found physiologically.
In their latest work, they created an image of 3.9 angstroms, which showed the receptor bound to two ligands: the GABA neurotransmitter or flumazenil. Flumazenil is a drug that blocks benzodiazepines, which are used as sedatives, anesthetics, anticonvulsants or as relaxation drugs. Flumazenil counteracts them.
“This work is an outstanding collection of data on the synaptic isoform of the human GABAA receptor,” said Graziano Pinna, University of Illinois, Chicago, who was not involved in the study. “It will help us understand why current drugs fail to exert pharmacological properties by acting at GABAA receptors and help anticipate future design of agents for the treatment of neurological and psychiatric disorders.”
Researchers at the Van Andel Research Institute (VARI) in Grand Rapids, Michigan, used Cryo-EM to visualize the interactions between a G-protein coupled receptor (GPCR) called rhodopsin bound to an inhibitory G protein. “Visualizing this complex resolves a missing chapter in the GPCR story by finally revealing how these two molecules interact in exquisite detail,” said H. Eric Xu, professor at Van Andel, and one of the study’s senior authors, in a statement. “Everything in biology is based on molecular interactions so the more we know about how the structures of these two molecules work together, the better position we are in to design improved medications with fewer undesired effects.”
GPCRs are embedded in the cell membrane and act as conduits between the cell and its external environment. They interact with G proteins and arrestins—a type of signaling molecule—to communicate messages to and from the cell regarding a range of cellular functions, including growth and immune responses.
Purdue University scientists published Cryo-EM structures in the journal Structure (Cell Press) of the Zika virus. “This is the most accurate picture we have of the virus so far,” said Michael Rossman, the Hanley Distinguished Professor of Biological Sciences at Purdue, in a statement. “These results will give us ways to efficiently design antiviral compounds and provide a basis for structure-based vaccine design.”
Zika viruses, part of a family of flaviviruses, are transmitted by mosquitos and ticks. Zika virus is correlated to babies born with abnormally small heads and sometimes smaller brains, called microcephaly, and can lead to Guillain-Barre syndrome in adults.
The researchers also looked at images of other flaviviruses such as Dengue, West Nile and Japanese Encephalitis virus, which have similar structures, but cause different diseases. Madhu Sevvana, a postdoctoral researcher and lead author of the paper, said in a statement, “We compared the surface properties of these viruses and observed differences in the landscape of the surface exposed residues. We found certain structural differences on the surface, which could be a starting point for further mutational analyses.”