Science Advances: Observation of the atomic structure of staphylococcal bacteriophages with cryo-electron microscopy

Researchers at the University of Alabama at Birmingham used cryo-electron microscopy to expose 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 structural knowledge is a critical link
between viral biology and potential therapeutic applications of viruses for suppressing bacterial infections.

Bacteriophage or "bacteriophage" is the term
used for viruses that infect bacteria.
The UAB researchers, led by Dr.
Terje Dokland, in collaboration with Dr.
Asma Hatoum-Aslan of the University of Illinois at Urbana-Champaign, described a full or partial atomic model
of 11 different structural proteins in the bacteriophage Andhra.
The study was published in the journal Science Advances
.

Andhra is a member of
the picovirus group.
Its host range is limited to Staphylococcus epidermidis (Staphylococcus epidermidis
).
This skin bacteria is mostly benign, but it is also a major cause
of infection in hospitalized medical devices.
Hatoum-Aslan, a bacteriophage biologist at the University of Illinois, said, "Dermatoviruses are rarely found in phage collections and remain understudied and underutilized
in therapeutic applications.
"

With the emergence of antibiotic resistance to Staphylococcus epidermidis and related pathogens Staphylococcus aureus, researchers have revived interest
in the potential use of bacteriophages to treat bacterial infections.
Dermatoviruses always kill the cells they infect, and after binding to the bacterial cell wall, they break through the cell wall through enzymes, penetrate the cell membrane, and inject viral DNA into the cell
.
They also have other properties that make them attractive candidates for therapeutic uses, including small genomes and the inability to transfer bacterial genes
between bacteria.

Understanding the protein structure of Andhra Pradesh and how these structures enable viruses to infect bacteria will make it possible to use genetic manipulation to produce custom phages suitable for specific purposes
.

"The structural basis of host-specificity between bacteriophages infected with Staphylococcus aureus and Staphylococcus epidermidis is still poorly understood," said
Dokland, UAB professor of microbiology and core director of UAB cryo-electron microscopy.
"With the current study, we have a better understanding of the structure and function of Andhra gene products and the determinants of host specificity, paving the way
for more rational design of custom phages for therapeutic applications.
" Our findings elucidate key features
of virion assembly, host recognition, and penetration.
”

Staphylococcal phages can generally infect a narrow range of bacteria, depending on the variable polymer
of phosphoric acid on the surface wall of different strains.
"This narrow host range is a double-edged sword: on the one hand, it allows phages to target only the specific pathogens that cause disease; On the other hand, this means that bacteriophages may need to be tailored to patients in each specific case," Dokland said
.

The general structure of Andhra is a 20-sided circular icosahedral capsid head containing a viral genome
.
The capsid is attached to
a short tail.
The tail is mainly responsible for binding to Staphylococcus epidermidis and breaking the cell wall
by enzymatic action.
Viral DNA is injected into the bacteria through the
tail.
The part of the tail includes the entrance from the capsid to the tail, as well as the stem, appendages, knobs, and tail tip
.

The 11 different proteins that make up each viral particle are found in multiple copies that come together together
.
For example, the capsid consists of 235 copies of each of two proteins, while the other nine virion proteins have copy numbers ranging from 2 to 72
.
In total, virions are made up of 645 protein fragments, including two copies of the 12th protein, whose structure was predicted using the protein structure prediction program AlphaFold
.

The atomic model described by Dokland, Hatoum-Aslan and co-first author N'Toia C.
Hawkins, Ph.
D.
, and James L.
Kizziah, Ph.
D.
, of the UAB Department of Microbiology, showed the structure of each protein—described in molecular language, such as α-helix, β-helix, β-chain, β-barrel, or β-prism
。 The researchers describe how each protein binds to other copies of the same protein type, such as the hexameric and pentameric faces that make up the capsid, and how each protein interacts
with different protein types adjacent to it.

Electron microscopes use a beam of accelerated electrons to illuminate an object, providing much higher resolution
than light microscopy.
Cryo-electron microscopy adds ultra-low temperature elements, making it particularly suitable for near-atomic structural resolution of large proteins, membrane proteins, or lipid-containing samples such as membrane-bound acceptors, as well as complexes
of several biomolecules.

Over the past eight years, new electron detectors have made cryo-electron microscopes a huge leap
in resolution over ordinary electron microscopes.
The key elements of this so-called cryo-electron microscope "resolution revolution" are:

• The water sample flash-frozen in liquid ethane cooled below minus 256 degrees Fahrenheit, and instead of ice crystals that destroy the sample and scatter the electron beam, the water freezes like window-like "glassy ice.
  "
  

• The sample is kept at ultra-low temperatures under the microscope and a low dose of electrons is used to avoid damage
  to the protein.
  

• Extremely fast direct electron detectors are capable of counting individual atoms at hundreds of frames per second, allowing the motion of the sample to be corrected
  in flight.
  

• Advanced computing techniques combine thousands of images to produce high-resolution three-dimensional structures
  .
  The graphics processing unit is used to process terabytes of data
  .
  

• The microscope stage where the sample is saved can also be tilted as the image is taken, creating a three-dimensional tomographic image, similar to a CT scan
  in a hospital.
  

UAB researchers' analysis of the structure of virions in Andhra Pradesh began with 230,714 particle images
.
Molecular reconstruction of the capsid, tail, distal tail, and tailtip began with 186,542, 159,489, 159,489, and 159,489 images, respectively
.
The resolution ranges from 3.
50 angstroms to 4.
90 angstroms
.