Cryo-electron microscopy by University of Alabama at Birmingham researchers has exposed the structure of a bacterial virus with unprecedented detail. That is the primary structure of a virus capable of infect Staphylococcus epidermidis, and high-resolution knowledge of structure is a key link between viral biology and potential therapeutic use of the virus to quell bacterial infections.

Bacteriophages or “phages” is the terms used for viruses that infect bacteria. The UAB researchers, led by Terje Dokland, Ph.D., in collaboration with Asma Hatoum-Aslan, Ph.D., on the University of Illinois Urbana-Champaign, have described atomic models for all or a 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 generally benign but additionally is a number one reason behind infections of indwelling medical devices. “Picoviruses are rarely present in phage collections and remain understudied and underused for therapeutic applications,” said Hatoum-Aslan, a phage biologist on the University of Illinois.

With emergence of antibiotic resistance in S. epidermidis and the related pathogen Staphylococcus aureus, researchers have renewed interest in potentially using bacteriophages to treat bacterial infections. Picoviruses at all times kill the cells they infect, after binding to the bacterial cell wall, enzymatically breaking through that wall, penetrating the cell membrane and injecting viral DNA into the cell. Additionally they produce other traits that make them attractive candidates for therapeutic use, including a small genome and an inability to transfer bacterial genes between bacteria.

Knowledge of protein structure in Andhra and understanding of how those structures allow the virus to contaminate a bacterium will make it possible to provide custom-made phages tailored to a selected purpose, using genetic manipulation.

The structural basis for host specificity between phages that infect S. aureus and S. epidermidis continues to be poorly understood. With the current study, we now have gained a greater understanding of the structures and functions of the Andhra gene products and the determinants of host specificity, paving the best way for a more rational design of custom phages for therapeutic applications. Our findings elucidate critical features for virion assembly, host recognition and penetration.”

Terje Dokland, professor of microbiology at UAB and director of the UAB Cryo-Electron Microscopy Core

Staphylococcal phages typically have a narrow range of bacteria they’ll infect, depending on the variable polymers of wall teichoic acid on the surface of various bacterial strains. “This narrow host range is a double-edged sword: On one hand, it allows the phages to focus on only the particular pathogen causing the disease; alternatively, it implies that the phage may should be tailored to the patient in each specific case,” Dokland said.

The overall structure of Andhra is a 20-faced, roundish icosahedral capsid head that incorporates the viral genome. The capsid is attached to a brief tail. The tail is basically accountable for binding to S. epidermidis and enzymatically breaking the cell wall. The viral DNA is injected into the bacterium through the tail. Segments of the tail include the portal from the capsid to the tail, and the stem, appendages, knob and tail tip.

The 11 different proteins that make up each virus particle are present in multiple copies that assemble together. As an example, the capsid is made from 235 copies each of two proteins, and the opposite nine virion proteins have copy numbers from two to 72. In total, the virion is made up of 645 protein pieces that include two copies of a twelfth protein, whose structure was predicted using the protein structure prediction program AlphaFold.

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 every protein -; as described in molecular language like alpha-helix, beta-helix, beta-strand, beta-barrel or beta-prism. The researchers have described how each protein binds to other copies of that very same protein type, comparable to to make up the hexameric and pentameric faces of the capsid, in addition to how each protein interacts with adjoining different protein types.

Electron microscopes use a beam of accelerated electrons to light up an object, providing much higher resolution than a lightweight microscope. Cryo-electron microscopy adds the element of super-cold temperatures, making it particularly useful for near-atomic structure resolution of larger proteins, membrane proteins or lipid-containing samples like membrane-bound receptors, and complexes of several biomolecules together.

Up to now eight years, latest electron detectors have created an incredible jump in resolution for cryo-electron microscopy over normal electron microscopy. Key elements of this so-called “resolution revolution” for cryo-electron microscopy are:

• Flash-freezing aqueous samples in liquid ethane cooled to below -256 degrees F. As a substitute of ice crystals that disrupt samples and scatter the electron beam, the water freezes to a window-like “vitreous ice.”
• The sample is kept at super-cold temperatures within the microscope, and a low dose of electrons is used to avoid damage to the proteins.
• Extremely fast direct electron detectors are capable of count individual atoms at tons of of frames per second, allowing sample movement to be corrected on the fly.
• Advanced computing merges hundreds of images to generate three-dimensional structures at high resolution. Graphics processing units are used to churn through terabytes of information.
• The microscope stage that holds the sample can be tilted as images are taken, allowing construction of a three-dimensional tomographic image, much like a CT scan on the hospital.

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

Source:

University of Alabama at Birmingham

Journal reference:

Hawkins, N.C., et al. (2022) Structure and host specificity of Staphylococcus epidermidis bacteriophage Andhra. Science Advances. doi.org/10.1126/sciadv.ade0459.