Computing methods for icosahedral and symmetry-mismatch reconstruction of viruses by cryo-electron microscopy

Three-dimensional(3 D)reconstruction of icosahedral viruses has played a crucial role in the development of cryoelectron microscopy single-particle reconstruction,with many cryo-electron microscopy techniques first established for structural studies of icosahedral viruses,owing to their high symmetry and large mass.This review summarizes the computational methods for icosahedral and symmetry-mismatch reconstruction of viruses,as well as the likely challenges and bottlenecks in virus reconstruction,such as symmetry mismatch reconstruction,contrast transformation function(CTF)correction,and particle distortion.

Effect of Electron Beam Orientation on Exit Wave Function via Simulation of Electron Dynamic Diffraction

Based on the electron dynamic diffraction, phase shift of the exit wave function vs misorientation of the incident electron beam from the exact zone axis has been calculated for the [001] oriented copper. The result shows that the peak of phase shift is the maximum at the atom position as the electron beam along the exact [001] zone axis, and the peak value of phase shift decreases as increases of the misorientation. At small misorientation, i.e. less than 5 degree, change of the phase shift is minimal. The peak value of phase shift decreases significantly when the incident beam deviates form the zone axis over 10 degree and the exit wave has a planar configuration as the misoriention angle arrives -17 degree. The effect of this phase shift characteristics on the information extracted from the hologram has also been considered.

Insights into varicella-zoster virus assembly from the B-and C-capsid at near-atomic resolution structures

Varicella-zoster is a highly communicable virus that can be transmitted through the airborne route.About one quarter of people are infected with this virus.Previous studies have described the structure of A-capsid and a blurred reconstruction of the C-capsid with icosahedral symmetry.In this study,we have determined the more precise detailed structures of the varicella-zoster virus(VZV)B-and C-capsid in icosahedral symmetry using a combination of block-based reconstruction and symmetry relaxation strategies.In addition,we are reporting structural details of the portal vertex reconstructions in five-fold symmetry and portal reconstructions in twelve-fold symmetry.The structures unveil the basis for the high thermal stability of the VZV capsid.The conformational flexibility of structural elements of the capsid plays a role in the assembly of the capsid and drives processes critical for the viral life cycle.The results of the study open up new avenues for the development of drugs against a highly prevalent and contagious pathogen.

Structures of the portal vertex reveal essential protein-protein interactions for Herpesvirus assembly and maturation

Dear Editor,Herpesviridae is a large family of double-stranded DNA(dsDNA)viruses that cause a variety of human diseases ranging from cold sores and chicken pox to congenital defects,blindness and cancer(Chayavichitsilp et al.,2009;Wang et al.,2018).In the past 70 years,substantial advances in our knowledge of the molecular biology of herpesviruses have led to insights into disease pathogenesis and management.However,the mechanism for capsid assembly that requires the ordered packing of about 4,000 protein subunits into the hexons,pentons and triplexes remains elusive.It is still a puzzle how initially identical subunits adopt both hexameric and pentameric conformations in the capsid and select the correct locations needed to form closed shells of the proper size.Biochemical and genetic studies have shown that the portal is involved in initiation of capsid assembly(Newcomb et al.,2005)and functions akin to a DNA-sensor coupling genome-packaging achieved by a genome-packaging machinery-“terminase complex”(Chen et al.,2020;Yunxiang Yang,2020)with icosahedral capsid maturation(Lokareddy et al.,2017).Structural investigations of the herpesvirus portal have proven challenging due to the small size of this dodecamer,which accounts for less than 1%of the total mass of the capsid protein layer and the technical difficulties involved in resolving non-icosahedral components of such large icosahedral viruses(diameter is∼1,250Å).Efforts of many investigators over two decades have made to reconstruct the cryo-electron microscopy(cryo-EM)structure of herpesvirus portal vertex and more recently near-atomic structures of two herpesvirus(herpes simplex virus type 1(HSV-1)and Kaposi’s sarcoma-associated herpesvirus(KSHV))portal vertices were reported(McElwee et al.,2018;Gong et al.,2019;Liu et al.,2019).

Structural changes of a bacteriophage upon DNA packaging and maturation

Dear Editor,Tailed,double-stranded DNA(dsDNA)bacteriophages,which belong to the order of Caudovirales,have a tail attached to a pentameric vertex of the icosahedral capsid shell(head)through a 12-fold portal(Johnson and Chiu,2007).The phages package genomic dsDNA into a round procapsid using the portal in complex with an ATP-dependent terminase complex as the motor.During packaging,the procapsid shell expands to a more angular intermediate to match the size of the viral genome(Guo et al.,2014).When the phage head is full,the portal detects internal pressure and conveys a signal from the inner capsid to the exterior,which triggers a sequence of events-the terminase complex cleaving mature DNA genome from a multi-genome concatemer,the release of the terminase complex from the portal,and the attachment of the tail complex-in the completion of phage assembly(Lander et al.,2006;Johnson and Chiu,2007).At the beginning of phage infection,the tail is responsible for receptor recognition,and the portal and tail act as a tunnel for DNA delivery into the host cytoplasm(Johnson and Chiu,2007).These mechanisms of DNA packaging and ejection may also be conserved in many other DNA viruses,including herpesvirus(Wang et al.,2020;Yang et al.,2020).

Protein interactions in the murine cytomegalovirus capsid revealed by cryoEM

Cytomegalovirus(CMV)is distinct among members of the Herpesviridae family for having the largest dsDNA genome(230 kb).Packaging of large dsDNA genome is known to give rise to a highly pressurized viral capsid,but molecular interactions conducive to the formation of CMV capsid resistant to pressurization have not been described.Here,we report a cryo electron microscopy(cryoEM)structure of the murine cytomegalovirus(MCMV)capsid at a 9.1Åresolution and describe the molecular interactions among the~3000 protein molecules in the MCMV capsid at the secondary structure level.Secondary structural elements are resolved to provide landmarks for correlating with results from sequence-based prediction and for structure-based homology modeling.The major capsid protein(MCP)upper domain(MCPud)containsα-helices andβ-sheets conserved with those in MCPud of herpes simplex virus type 1(HSV-1),with the largest differences identifi ed as a“saddle loop”region,located at the tip of MCPud and involved in interaction with the smallest capsid protein(SCP).Interactions among the bacteriophage HK97-like fl oor domain of MCP,the middle domain of MCP,the hook and clamp domains of the triplex proteins(hoop and clamp domains of TRI-1 and clamp domain of TRI-2)contribute to the formation of a mature capsid.These results offer a framework for understanding how cytomegalovirus uses various secondary structural elements of its capsid proteins to build a robust capsid for packaging its large dsDNA genome inside and for attach-ing unique functional tegument proteins outside.

Structural and functional analysis of a potent human neutralizing antibody against enterovirus A71

Enterovirus A71(EV-A71) causes major outbreaks of hand,foot,and mouth disease(HFMD) in many countries,most frequently affecting children,and a small proportion of cases may lead to death.Currently,no vaccine is available in most endemic regions,and no licenced treatments for EV-A71 infection are available.Here,we characterize a human monoclonal antibody(Hu MAb),E1,by screening a Fab antibody phage library derived from patients who recovered from EV-A71 infection.E1 exhibits strong neutralizing activity against EV-A71 virus in cells.The cryo-electron microscopy(cryo-EM) structures of the EV-A71 virion in complex with E1 Fab fragments demonstrated that E1 recognized an epitope formed by residues in the BC and HI loops of VP1.In a mouse model,E1 effectively protected against lethal EV-A71 challenge in both prophylactic and therapeutic treatment.In particular,E1 significantly reduces virus titers and muscle damage.E1 might represent a potential adjunct to EV-A71 treatment.