scholarly journals Cryo-EM analysis of a viral portal protein in situ reveals a switch in the DNA tunnel

2019 ◽  
Author(s):  
Oliver W. Bayfield ◽  
Alasdair C. Steven ◽  
Alfred A. Antson
Keyword(s):  
Surface Properties ◽  
Dynamic Processes ◽  
Viral Capsid ◽  
Capsid Assembly ◽  
Genome Packaging ◽  
Dna Viruses ◽  
Portal Protein ◽  
Double Stranded Dna ◽  
Protein Functions

The portal protein is a key component of many double-stranded DNA viruses, governing capsid assembly and genome packaging. Twelve subunits of the portal protein form a ring with a central tunnel, through which DNA is translocated into the capsid. It is unknown how the portal protein functions as a gatekeeper, preventing DNA slippage, whilst allowing its passage into the capsid through its central tunnel, and how these processes can be controlled by capsid and motor proteins. A cryo-EM structure of a portal protein, determined in situ for immature capsids of thermostable bacteriophage P23-45, suggests how domain adjustments can be coupled with a switching of properties of the DNA tunnel. Of particular note is an inversion of the conformation of portal loops which define the tunnel’s constriction, accompanied by a switching of surface properties from hydrophobic to hydrophilic. These observations indicate how translocation of DNA into the viral capsid can be modulated by changes in the properties and size of the central tunnel and how the changing pattern of protein–capsid interactions across a symmetry-mismatched interface can facilitate these dynamic processes.

scholarly journals Cryo-EM structure in situ reveals a molecular switch that safeguards virus against genome loss

eLife ◽  
2020 ◽  
Vol 9 ◽  
Author(s):  
Oliver W Bayfield ◽  
Alasdair C Steven ◽  
Alfred A Antson
Keyword(s):  
Protein Interactions ◽  
Bound State ◽  
Molecular Switch ◽  
Protein Protein Interactions ◽  
Genome Packaging ◽  
Dna Viruses ◽  
Portal Protein ◽  
Double Stranded Dna ◽  
Protein Functions

The portal protein is a key component of many double-stranded DNA viruses, governing capsid assembly and genome packaging. Twelve subunits of the portal protein define a tunnel, through which DNA is translocated into the capsid. It is unknown how the portal protein functions as a gatekeeper, preventing DNA slippage, whilst allowing its passage into the capsid, and how these processes are controlled. A cryo-EM structure of the portal protein of thermostable virus P23-45, determined in situ in its procapsid-bound state, indicates a mechanism that naturally safeguards the virus against genome loss. This occurs via an inversion of the conformation of the loops that define the constriction in the central tunnel, accompanied by a hydrophilic–hydrophobic switch. The structure also shows how translocation of DNA into the capsid could be modulated by a changing mode of protein–protein interactions between portal and capsid, across a symmetry-mismatched interface.

scholarly journals Adintoviruses: A Proposed Animal-Tropic Family of Midsize Eukaryotic Linear dsDNA (MELD) Viruses

2020 ◽  
Author(s):  
Gabriel J Starrett ◽  
Michael J Tisza ◽  
Nicole L Welch ◽  
Anna K Belford ◽  
Alberto Peretti ◽  
...  
Keyword(s):  
Genome Packaging ◽  
Metagenomic Sequence ◽  
Dna Viruses ◽  
Diverse Range ◽  
Integrase Gene ◽  
Data Mining Approach ◽  
Double Stranded Dna ◽  
Wide Range ◽  
Linear Dsdna ◽  
Dsdna Viruses

Abstract Polintons (also known as Mavericks) were initially identified as a widespread class of eukaryotic transposons named for their hallmark type B DNA polymerase and retrovirus-like integrase genes. It has since been recognized that many polintons encode possible capsid proteins and viral genome-packaging ATPases similar to those of a diverse range of double-stranded DNA (dsDNA) viruses. This supports the inference that at least some polintons are actually viruses capable of cell-to-cell spread. At present, there are no polinton-associated capsid protein genes annotated in public sequence databases. To rectify this deficiency, we used a data-mining approach to investigate the distribution and gene content of polinton-like elements and related DNA viruses in animal genomic and metagenomic sequence datasets. The results define a discrete family-like clade of viruses with two genus-level divisions. We propose the family name Adintoviridae, connoting similarities to adenovirus virion proteins and the presence of a retrovirus-like integrase gene. Although adintovirus-class PolB sequences were detected in datasets for fungi and various unicellular eukaryotes, sequences resembling adintovirus virion proteins and accessory genes appear to be restricted to animals. Degraded adintovirus sequences are endogenized into the germlines of a wide range of animals, including humans.

scholarly journals Principles for enhancing virus capsid capacity and stability from a thermophilic virus capsid structure

2019 ◽  
Vol 10 (1) ◽  
Author(s):  
Nicholas P. Stone ◽  
Gabriel Demo ◽  
Emily Agnello ◽  
Brian A. Kelch
Keyword(s):  
Major Capsid Protein ◽  
Extreme Environment ◽  
Structural Basis ◽  
Genome Packaging ◽  
Virus Capsid ◽  
Dna Viruses ◽  
Double Stranded Dna ◽  
High Internal Pressure ◽  
Extracellular Environment ◽  
Catch Bonds

Abstract The capsids of double-stranded DNA viruses protect the viral genome from the harsh extracellular environment, while maintaining stability against the high internal pressure of packaged DNA. To elucidate how capsids maintain stability in an extreme environment, we use cryoelectron microscopy to determine the capsid structure of thermostable phage P74-26 to 2.8-Å resolution. We find P74-26 capsids exhibit an overall architecture very similar to those of other tailed bacteriophages, allowing us to directly compare structures to derive the structural basis for enhanced stability. Our structure reveals lasso-like interactions that appear to function like catch bonds. This architecture allows the capsid to expand during genome packaging, yet maintain structural stability. The P74-26 capsid has T = 7 geometry despite being twice as large as mesophilic homologs. Capsid capacity is increased with a larger, flatter major capsid protein. Given these results, we predict decreased icosahedral complexity (i.e. T ≤ 7) leads to a more stable capsid assembly.

scholarly journals Cryo-EM structure and in vitro DNA packaging of a thermophilic virus with supersized T=7 capsids

2019 ◽  
Vol 116 (9) ◽  
pp. 3556-3561 ◽  
Author(s):  
Oliver W. Bayfield ◽  
Evgeny Klimuk ◽  
Dennis C. Winkler ◽  
Emma L. Hesketh ◽  
Maria Chechik ◽  
...  
Keyword(s):  
Conformational Changes ◽  
Structural Integrity ◽  
Thermus Thermophilus ◽  
Dna Packaging ◽  
Dna Viruses ◽  
Portal Protein ◽  
Single Vertex ◽  
Double Stranded Dna ◽  
Β Sheets

Double-stranded DNA viruses, including bacteriophages and herpesviruses, package their genomes into preformed capsids, using ATP-driven motors. Seeking to advance structural and mechanistic understanding, we established in vitro packaging for a thermostable bacteriophage, P23-45 of Thermus thermophilus. Both the unexpanded procapsid and the expanded mature capsid can package DNA in the presence of packaging ATPase over the 20 °C to 70 °C temperature range, with optimum activity at 50 °C to 65 °C. Cryo-EM reconstructions for the mature and immature capsids at 3.7-Å and 4.4-Å resolution, respectively, reveal conformational changes during capsid expansion. Capsomer interactions in the expanded capsid are reinforced by formation of intersubunit β-sheets with N-terminal segments of auxiliary protein trimers. Unexpectedly, the capsid has T=7 quasi-symmetry, despite the P23-45 genome being twice as large as those of known T=7 phages, in which the DNA is compacted to near-crystalline density. Our data explain this anomaly, showing how the canonical HK97 fold has adapted to double the volume of the capsid, while maintaining its structural integrity. Reconstructions of the procapsid and the expanded capsid defined the structure of the single vertex containing the portal protein. Together with a 1.95-Å resolution crystal structure of the portal protein and DNA packaging assays, these reconstructions indicate that capsid expansion affects the conformation of the portal protein, while still allowing DNA to be packaged. These observations suggest a mechanism by which structural events inside the capsid can be communicated to the outside.

scholarly journals The Ins and Outs of Herpesviral Capsids: Divergent Structures and Assembly Mechanisms across the Three Subfamilies

Viruses ◽  
2021 ◽  
Vol 13 (10) ◽  
pp. 1913
Author(s):  
Elizabeth B. Draganova ◽  
Jonathan Valentin ◽  
Ekaterina E. Heldwein
Keyword(s):  
Capsid Protein ◽  
Severe Disease ◽  
Human Herpesvirus ◽  
Dna Viruses ◽  
Double Stranded Dna ◽  
Cryo Electron Microscopy ◽  
Protein Functions ◽  
Attractive Therapeutic Target ◽  
Key Steps ◽  
Human Herpesviruses

Human herpesviruses, classified into three subfamilies, are double-stranded DNA viruses that establish lifelong latent infections within most of the world’s population and can cause severe disease, especially in immunocompromised people. There is no cure, and current preventative and therapeutic options are limited. Therefore, understanding the biology of these viruses is essential for finding new ways to stop them. Capsids play a central role in herpesvirus biology. They are sophisticated vehicles that shelter the pressurized double-stranded-DNA genomes while ensuring their delivery to defined cellular destinations on the way in and out of the host cell. Moreover, the importance of capsids for multiple key steps in the replication cycle makes their assembly an attractive therapeutic target. Recent cryo-electron microscopy reconstructions of capsids from all three subfamilies of human herpesviruses revealed not only conserved features but also remarkable structural differences. Furthermore, capsid assembly studies have suggested subfamily-specific roles of viral capsid protein homologs. In this review, we compare capsid structures, assembly mechanisms, and capsid protein functions across human herpesvirus subfamilies, highlighting the differences.

scholarly journals Principles for enhancing virus capsid capacity and stability from a thermophilic virus capsid structure

2018 ◽  
Author(s):  
Nicholas P. Stone ◽  
Gabriel Demo ◽  
Emily Agnello ◽  
Brian A. Kelch
Keyword(s):  
Major Capsid Protein ◽  
Extreme Environment ◽  
Structural Basis ◽  
Genome Packaging ◽  
Virus Capsid ◽  
Dna Viruses ◽  
Double Stranded Dna ◽  
High Internal Pressure ◽  
Extracellular Environment ◽  
Catch Bonds

SUMMARYThe capsids of double-stranded DNA viruses protect the viral genome from the harsh extracellular environment, while maintaining stability against the high internal pressure of packaged DNA. To elucidate how capsids maintain stability in an extreme environment, we used cryoelectron microscopy to determine the capsid structure of the thermostable phage P74-26 to 2.8-Å resolution. We find the P74-26 capsid exhibits an overall architecture that is very similar to those of other tailed bacteriophages, allowing us to directly compare structures to derive the structural basis for enhanced stability. Our structure reveals ‘lasso’-like interactions that appear to function like catch bonds. This architecture allows the capsid to expand during genome packaging, yet maintain structural stability. The P74-26 capsid has T=7 geometry despite being twice as large as mesophilic homologs. Capsid capacity is increased through a novel mechanism with a larger, flatter major capsid protein. Our results suggest that decreased icosahedral complexity (i.e. lower T number) leads to a more stable capsid assembly.

scholarly journals Portal Protein: The Orchestrator of Capsid Assembly for the dsDNA Tailed Bacteriophages and Herpesviruses

2019 ◽  
Vol 6 (1) ◽  
pp. 141-160 ◽  
Author(s):  
Corynne L. Dedeo ◽  
Gino Cingolani ◽  
Carolyn M. Teschke
Keyword(s):  
Drug Target ◽  
Sequence Similarity ◽  
Antiviral Drug ◽  
Viral Assembly ◽  
Dna Packaging ◽  
Capsid Assembly ◽  
Portal Protein ◽  
Ring Assembly ◽  
Double Stranded Dna

Tailed, double-stranded DNA bacteriophages provide a well-characterized model system for the study of viral assembly, especially for herpesviruses and adenoviruses. A wealth of genetic, structural, and biochemical work has allowed for the development of assembly models and an understanding of the DNA packaging process. The portal complex is an essential player in all aspects of bacteriophage and herpesvirus assembly. Despite having low sequence similarity, portal structures across bacteriophages share the portal fold and maintain a conserved function. Due to their dynamic role, portal proteins are surprisingly plastic, and their conformations change for each stage of assembly. Because the maturation process is dependent on the portal protein, researchers have been working to validate this protein as a potential antiviral drug target. Here we review recent work on the role of portal complexes in capsid assembly, including DNA packaging, as well as portal ring assembly and incorporation and analysis of portal structures.

scholarly journals ATP serves as a nucleotide switch coupling the genome maturation and packaging motor complexes of a virus assembly machine

2020 ◽  
Vol 48 (9) ◽  
pp. 5006-5015
Author(s):  
Qin Yang ◽  
Carlos E Catalano
Keyword(s):  
Atp Hydrolysis ◽  
Virus Assembly ◽  
Genome Packaging ◽  
Dna Viruses ◽  
Motor Complex ◽  
Double Stranded Dna ◽  
Tightly Coupled ◽  
Switch Mechanism ◽  
Genome Maturation ◽  
Dsdna Viruses

Abstract The assembly of double-stranded DNA viruses, from phages to herpesviruses, is strongly conserved. Terminase enzymes processively excise and package monomeric genomes from a concatemeric DNA substrate. The enzymes cycle between a stable maturation complex that introduces site-specific nicks into the duplex and a dynamic motor complex that rapidly translocates DNA into a procapsid shell, fueled by ATP hydrolysis. These tightly coupled reactions are catalyzed by terminase assembled into two functionally distinct nucleoprotein complexes; the maturation complex and the packaging motor complex, respectively. We describe the effects of nucleotides on the assembly of a catalytically competent maturation complex on viral DNA, their effect on maturation complex stability and their requirement for the transition to active packaging motor complex. ATP plays a major role in regulating all of these activities and may serve as a ‘nucleotide switch’ that mediates transitions between the two complexes during processive genome packaging. These biological processes are recapitulated in all of the dsDNA viruses that package monomeric genomes from concatemeric DNA substrates and the nucleotide switch mechanism may have broad biological implications with respect to virus assembly mechanisms.

scholarly journals Portal protein functions akin to a DNA-sensor that couples genome-packaging to icosahedral capsid maturation

2017 ◽  
Vol 8 (1) ◽  
Author(s):  
Ravi K. Lokareddy ◽  
Rajeshwer S. Sankhala ◽  
Ankoor Roy ◽  
Pavel V. Afonine ◽  
Tina Motwani ◽  
...  
Keyword(s):  
Dna Sensor ◽  
Genome Packaging ◽  
Portal Protein ◽  
Protein Functions ◽  
Capsid Maturation ◽  
Icosahedral Capsid

scholarly journals Adintoviruses: An Animal-Tropic Family of Midsize Eukaryotic Linear dsDNA (MELD) Viruses

2019 ◽  
Author(s):  
Gabriel J. Starrett ◽  
Michael J. Tisza ◽  
Nicole L. Welch ◽  
Anna K. Belford ◽  
Alberto Peretti ◽  
...  
Keyword(s):  
Genome Packaging ◽  
Metagenomic Sequence ◽  
Dna Viruses ◽  
Diverse Range ◽  
Data Mining Approach ◽  
Double Stranded Dna ◽  
Wide Range ◽  
The Family ◽  
Linear Dsdna ◽  
Dsdna Viruses

AbstractPolintons (also known as Mavericks) were initially identified as a widespread class of eukaryotic transposons named for their hallmark type B DNA polymerase and retrovirus-like integrase genes. It has since been recognized that many polintons encode possible capsid proteins and viral genome-packaging ATPases similar to those of a diverse range of double-stranded DNA (dsDNA) viruses. This supports the inference that at least some polintons are viruses that remain capable of cell-to-cell spread. At present, there are no polinton-associated capsid protein genes annotated in public sequence databases. To rectify this deficiency, we used a data-mining approach to investigate the distribution and gene content of polinton-like elements and related DNA viruses in animal genomic and metagenomic sequence datasets. The results define a discrete family-like clade of animal-specific viruses with two genus-level divisions. We suggest the family name Adintoviridae, connoting similarities to adenovirus virion proteins and the presence of a retrovirus-like integrase gene. Although adintovirus-class PolB sequences were detected in datasets for fungi and various unicellular eukaryotes, sequences resembling adintovirus virion proteins and accessory genes appear to be restricted to animals. Degraded adintovirus sequences are endogenized into the germlines of a wide range of animals, including humans.

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