Atomic structure of a staphylococcus bacterium

The Andhra phage

Image: The Andhra phage
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Photo credits: UAB, Dokland laboratory

BIRMINGHAM, Alabama. – Cryo-electron microscopy by researchers at the University of Alabama at Birmingham has revealed the structure of a bacterial virus in unprecedented detail. This is the first structure of a virus capable of infecting Staphylococcus epidermidis, and high-resolution knowledge of the structure is a key link between viral biology and the potential therapeutic use of the virus to suppress bacterial infections.

Bacteriophage or “phages” are the names for viruses that infect bacteria. The UAB researchers, led by Terje Dokland, Ph.D., in collaboration with Asma Hatoum-Aslan, Ph.D. at the University of Illinois Urbana-Champaign, have described atomic models for all or part of 11 different structural proteins in Phage Andhra. The study is published in Science Advances.

Andhra is a member of the Picovirus group. Its host range is restricted to S. epidermidis. This skin bacterium is mostly benign, but it is also a major cause of infection of medical devices embedded in the skin. “Picoviruses are rarely found in phage collections and remain understudied and underutilized for therapeutic applications,” said Hatoum-Aslan, a phage biologist at the University of Illinois.

With the emergence of antibiotic resistance in S. epidermidis and the related pathogen Staphylococcus aureus, researchers have renewed interest in the potential use of bacteriophage to treat bacterial infections. Picoviruses always kill the cells they infect after attaching to the bacterial cell wall, enzymatically breaking through that wall, penetrating the cell membrane and injecting viral DNA into the cell. They also have other characteristics that make them attractive candidates for therapeutic use, including a small genome and an inability to transfer bacterial genes between bacteria.

Knowing the protein structure in Andhra and understanding how these structures allow the virus to infect a bacterium will make it possible to use genetic manipulation to produce bespoke phages tailored for a specific purpose.

“The structural basis for host specificity between phages infecting S. aureus and S. epidermidis is still poorly understood,” said Dokland, professor of microbiology at UAB and director of the UAB Cryo-Electron Microscopy Core. “With the present study, we have gained a better understanding of the structures and functions of the Andhra gene products and the determinants of host specificity, paving the way for a more rational design of tailored phages for therapeutic applications. Our results highlight critical features for virion assembly, host recognition, and penetration.”

Staphylococcal phages typically have a narrow range of bacteria to infect, depending on the variable polymers of wall teichoic acid on the surface of different bacterial strains. “This narrow host range is a double-edged sword: on the one hand, it allows the phage to target only the specific pathogen; on the other hand, it means that in each individual case, the phage may have to be tailored to the patient,” Dokland said.

The general structure of Andhra is a 20-faced, rounded icosahedral capsid head containing the viral genome. The capsid is attached to a short tail. The tail is primarily responsible for binding to S. epidermidis and enzymatically disrupting the cell wall. The viral DNA is injected into the bacterium through the tail. The segments of the tail include the portal from the capsid to the tail, as well as the stalk, appendages, knob, and tail tip.

The 11 different proteins that make up each virus particle are in multiple copies that join together. For example, the capsid consists of 235 copies each of two proteins, and the other nine virion proteins have copy numbers from two to 72. In total, the virion consists of 645 pieces of protein, including two copies of a 12th protein, whose structure using the protein structure prediction program AlphaFold was predicted.

The atomic models described by Dokland, Hatoum-Aslan, and co-first authors N’Toia C. Hawkins, Ph.D., and James L. Kizziah, Ph.D., UAB Department of Microbiology show the structures for each protein—how described in molecular language as alpha helix, beta helix, beta strand, beta barrel or beta prism. Researchers have described how each protein binds to other copies of the same protein type to form, for example, the hexameric and pentameric faces of the capsid, and how each protein interacts with neighboring different protein types.

Electron microscopes use a beam of accelerated electrons to illuminate an object and offer much higher resolution than a light microscope. Cryo-electron microscopy adds the element of supercold temperatures, making it particularly useful for near-atomic structural resolution of larger proteins, membrane proteins, or lipid-containing samples such as membrane-bound receptors and complexes composed of multiple biomolecules.

In the last eight years, new electron detectors have created an enormous leap in resolution for cryo-electron microscopy compared to normal electron microscopy. Key elements of this so-called “resolution revolution” for cryo-electron microscopy are:

  • Flash freezing of aqueous samples in liquid ethane chilled to below -256 degrees F. Instead of ice crystals destroying the samples and scattering the electron beam, the water freezes into window-like “glassy ice.”
  • The sample is kept at super-cold temperatures in the microscope, and a low dose of electrons is used to avoid damaging the proteins.
  • Extremely fast direct electron detectors are capable of counting single atoms at hundreds of frames per second, allowing for sample motion correction on the fly.
  • Advanced computing stitches thousands of images together to create high-resolution, three-dimensional structures. GPUs are used to process terabytes of data.
  • The microscope stage holding the sample can also be tilted during acquisition, allowing the creation of a three-dimensional tomographic image, similar to a hospital CT scan.

Analysis of Andhra virion structure by the UAB researchers started with 230,714 particle images. Molecular reconstruction of the capsid, tail, distal tail, and tail tip started with 186,542, 159,489, 159,489, and 159,489 images, respectively. The resolution ranged from 3.50 to 4.90 angstroms.

Support for the study, Structure and Host Specificity of the Staphylococcus epidermidis Andhra Bacteriophage, came from the National Institutes of Health Phage Therapy Grant R21 AI156636.

The UAB Department of Microbiology is part of the Marnix E. Heersink School of Medicine.

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