Available online at www.sciencedirect.com

ScienceDirect

All-atom simulations

Jodi A Hadden and Juan R Perilla

The constant threat of viral disease can be combated by the All-atom molecular dynamics (MD) simulations, often

development of novel vaccines and therapeutics designed to referred to as the ‘computational microscope’, remain

disrupt key features of virus structure or infection cycle the only method capable of elucidating the dynamical

processes. Such development relies on high-resolution properties of biomolecules within their physiological



characterization of and their dynamical behaviors, environments at full chemical resolution [1 ]. Modern

which are often challenging to obtain solely by experiment. In supercomputers and MD simulation codes enable

response, all-atom molecular dynamics simulations are widely researchers to investigate the nuances of virus structure

leveraged to study the structural components of viruses, and dynamics, even for very large components of virus

leading to some of the largest simulation endeavors undertaken structure [2].

to date. The present work reviews exemplary all-atom

simulation work on viruses, as well as progress toward Most all-atom MD simulation studies of viruses have

simulating entire virions. focused on their protein-based components, particularly

envelope proteins, , , and accessory pro-

Address teins or enzymes. Some work has leveraged MD simula-

Department of Chemistry and Biochemistry, University of Delaware, tions to construct complete atomistic models of viral

Newark, DE 19716, United States proteins or their assemblies by integrating experimental

data. Other work has taken advantage of MD simulations

Corresponding author: Hadden, Jodi A ([email protected])

to characterize, under native conditions, the chemical-

physical properties of such structures through analysis of

Current Opinion in Virology 2018, 31:82–91 their dynamics and influence on surrounding solvent. The

overarching goals of virus simulation endeavors are to

This review comes from a themed issue on Virus structure and

expression reveal critical aspects of virus structure or mechanistic

insights into their function that can be targeted for novel

Edited by Kay Gru¨ newald and Thomas Krey

disease treatments. Here, exemplary all-atom simulations

For a complete overview see the Issue and the Editorial

of viruses are briefly reviewed, and progress toward sim-

Available online 1st September 2018

ulating entire virions at atomistic detail is discussed.

https://doi.org/10.1016/j.coviro.2018.08.007

1879-6257/ã 2018 Elsevier B.V. All rights reserved. Envelope proteins

The first stage of viral infection is cell entry. Often, this

process begins with adhesion of the virus to the host cell

via binding. In enveloped viruses, receptor

binding is mediated by membrane-embedded surface

proteins, which typically also represent the major anti-

Introduction gens of the virus and may participate in virus–host mem-

brane fusion. Other envelope proteins play a role in viral

Viruses are a constant threat to human health, as well as egress. The motivation to elucidate the molecular deter-

the health of our domesticated animals and agriculture. minants of receptor recognition, as well as the antigenic

Often, viral infections are challenging to treat due to and fusogenic properties of envelope proteins, have ren-

limited therapeutic options. Although vaccines are effec- dered them of great interest for study with MD

tive at preventing or reducing the severity of viral disease, simulations.

protective vaccines against fewer than 20 different viruses

are currently available (WHO). Further, the high muta- A broadly studied protein is the influenza A

tion rates of some viruses necessitate that vaccines be (HA) trimer (Figure 1a), which is respon-

updated annually, or that new antiviral treatments be sible for cellular adhesion and membrane fusion. Much

constantly realized to continue disease management in work on HA has been aimed at understanding its speci-

chronically infected individuals. ficity for host cell receptor glycans, which is an important

factor in the avian-human transmission barrier [6,7].

The rational development of novel prophylactic and Characterization of receptor binding may support the

therapeutic interventions against viral pathogens depends development of adhesion-inhibiting drugs.

on detailed knowledge of virus structure, infection cycle

processes, and the conformational dynamics that link All-atom MD simulations have been applied to numerous

structures to their functional roles in these processes. crystal complexes of HAs with receptor analogues to

Current Opinion in Virology 2018, 31:82–91 www.sciencedirect.com

All-atom simulations of viruses Hadden and Perilla 83

Figure 1

(a)NA (b) (c) RNA

Env M1 M2

RNA

RNA Vpu RT

capsid

HA envelope envelope matrix

Current Opinion in Virology

Schematic illustrations of viruses whose protein components have been heavily studied using all-atom MD simulations. (a) A virus virion,

average diameter of 120 nm [3]. Hemagglutinin (HA), neuraminidase (NA), and the matrix protein 2 (M2) are embedded within the viral

envelope, which surrounds the matrix protein 1 (M1) assembly and encloses the RNA. (b) Mature HIV-1 virion, average diameter of 145 nm [4]. The

envelope protein () and viroporin (Vpu) are embedded within the viral envelope, which surrounds the matrix protein assembly and encloses the



RNA-containing capsid. The virus also carries copies of a protease and (RT). (c) STMV virion, diameter of 17 nm [5 ]. STMV

is a non-enveloped virus, composed only of the RNA-containing capsid.

characterize binding mode dynamics, as well as to study virions. Several compounds that mimic the native sub-

recognition and specificity based on bound receptor con- strate of NA and bind to inhibit its action have been

formations and interaction profiles [8–12]. Recent model- licensed for use as antiviral drugs for the treatment of

ing and simulation work has also revealed that HA recep- influenza A (i.e. tamivir and ) [33]. The majority

tor specificity may depend on extended glycan structure of simulation work on NA has focused on exploiting it as a

and the ability to accommodate bidentate binding to drug target. In some cases, all-atom simulations have

biantennary glycans [13,14]. MD simulations have been been employed to characterize the conformational

further employed to evaluate the effects of HA dynamics of NA from the perspective of designing novel

that alter receptor binding and specificity [15,16] and to drug compounds or elucidating the determinants of

study HA- recognition and escape [17,18]. ligand recognition and inhibition [34–37]. In other cases,

simulations of NA have been used as part of computa-

Notably, HA is glycosylated, with N-glycans contributing tional screening protocols to discover new druggable hot

around 20% of its molecular weight [6]. All-atom simula- spots or inhibitor leads [38–40]. Additional studies have

tions of glycosylated HAs have been used to probe the provided insights into the mechanisms of drug resistance

influence of on receptor binding [19], to [41–43], the role of calcium in NA stability and drug

provide insight into accessibility of glycan-remodeling binding [44], and the structural basis for increased viru-



enzymes [20], and to characterize interactions between lence of NA stalk-deletion mutants [45 ].

HA and lung surfactant protein D, an innate immune

lectin that neutralizes influenza A via attachment to HA Another well-studied envelope protein is the human



glycans [21 ]. immunodeficiency virus type 1 (HIV-1) envelope (Env)

trimer (Figures 1b and 2 d), which mediates cellular

HA initiates fusion of the viral envelope and host mem- adhesion and membrane fusion. All-atom MD simula-

branes following cell adhesion and endocytosis, when tions have been employed to characterize the conforma-

endosomal acidification triggers it to undergo a confor- tional properties of Env, particularly its V3 variable

mational change. The mechanism of HA-mediated mem- domain, a critical determinant of CD4 receptor binding

brane fusion has been extensively studied with simula- [46,47]. Other simulation work on Env has focused on

tion approaches, including characterization of fusion identifying correlated motions that may underlie alloste-

dynamics, interactions with the host membrane, ric communication networks within the protein [47–50].

and structural effects of reduced pH [22–32]. The transmembrane domain and fusion peptide of Env

have also been extensively investigated with MD simula-

The secondary surface of the influenza A virus is tions to explore conformational variability and membrane

the neuraminidase (NA) tetramer (Figure 1a), which interactions, providing insights into the mechanism of

cleaves host cell glycans to allow escape of progeny membrane fusion [51–56].

www.sciencedirect.com Current Opinion in Virology 2018, 31:82–91

84 Virus structure and expression

Importantly, Env is heavily glycosylated, with N -glycans been aimed at characterizing drug binding and channel

accounting for roughly 50% of its molecular weight [57]. inhibition [84,85,90], probing the mechanisms of drug

These glycans form a shield around Env, cloaking it from resistance [91,92], and designing new inhibitors against

the host , and may also influence the drug-resistant mutants [93,94].

binding affinity of CD4. The glycan shield is of great

interest as a target for broadly- and Other viroporins that have been examined using all-atom

vaccine design [57], and can only be accurately described MD simulations include the U (Vpu) of HIV-

with simulations that encompass chemical detail. Several 1 (Figures 1b and 2 a). Various early modeling and

all-atom MD studies have investigated the effects of simulation studies of Vpu [95] led to evaluation of its

glycosylation on the conformation and dynamics of Env preferred oligomeric state and all-atom investigation of its

variable domains V1, V2, and V3, as well as implications dynamics and channel activity [96]. All-atom simulations

for receptor and antibody binding [58–60]. More recent were also applied to develop a model for the Paramecium

work has used computational modeling and simulation to bursaria chlorella virus type 1 (PBCV-1) Kcv potassium

produce fully glycosylated Env trimers, with the goals of channel to probe structure–function relationships [97,98]

characterizing the dynamical behavior of the glycan and elucidate its mechanism of ion transport [99]. More

shield and determining its impact on antibody elicitation, recently, all-atom simulations were employed to deter-



recognition, and binding [50,61 ,62,63]. mine the structure of the virus (HCV) p7

viroporin monomer [100] and to model various channel

Beyond influenza A and HIV-1, all-atom MD simulations oligomer states, enabling characterization of their dynam-



have also been applied to study the envelope proteins of ics and conductance properties [101 ,102,103,104,105].

several other viruses. Such work has included investiga-

tions of the effects of varying pH on the dengue virus [64– Capsids

69] and vesicular stomatitis virus (VSV) [70] envelope Many viruses package their genome within a remarkable

proteins, structure and host membrane interactions for protein shell called a capsid. Often, capsids play key

the Ebola virus [71–73] and virus (HSV) functional roles in delivering the viral genome to the host

[74] envelope protein fusion , solution dynamics cell nucleus, rendering them of great interest as drug

of human parainfluenza virus type 3 (HPIV-3) hemagglu- targets. Capsids generally represent the largest protein

tinin-neuraminidase [75], and conformational flexibility components of virus structures, and simulations of intact

in the hepatitis C virus (HCV) E2 envelope protein capsids have accounted for the most substantial all-atom

[76,77]. virus simulations yet published [106,2]. Work by Perilla

et al. has clearly demonstrated the need for chemical

Viroporins detail to facilitate accurate simulation studies of virus



Enveloped viruses can encode another class of mem- capsids as drug targets [107 ].

brane-embedded proteins, called viroporins, that play

diverse roles in their respective viral infection cycles. The first all-atom simulation of a complete virus capsid

Viroporins are small, hydrophobic proteins that oligomer- was based on satellite tobacco mosaic virus (STMV,

ize within the envelope to form hydrophilic channels, Figures 1c and 2 g) and encompassed a landmark particle



capable of transporting ionic species and small molecules count of 1 million atoms with solvent [108 ]. Both an

[78]. The essential activities of these channels, their empty and genome-containing capsid were investigated

druggability, and potential as models to understand trans- over 10–13 ns. With packaged RNA, the simulated capsid

port mechanisms within the human cell, have rendered maintained structural integrity and exhibited only minor

them of long-standing interest for study with MD deviations from icosahedral symmetry. Without RNA, the

simulations. simulated capsid rapidly broke symmetry and began to

implode. The results of the work strongly suggest that

The most widely studied viroporin, from a computational RNA is responsible for nucleating assembly of STMV

perspective, is the influenza A matrix protein 2 (M2) ion virions.

channel (Figure 1a), which transports protons across the

viral envelope following endosomal acidification to pre- Prior to work on STMV, simulations of complete capsids

pare the particle for host membrane fusion and subse- were approximated using rotational symmetry boundary

quent genome release [79]. All-atom MD simulations conditions, which exploit the icosahedral symmetry of

have been broadly employed to study M2 channel acti- capsid structures to model, at least in effect, a complete

vation, gating, and proton permeation [80–86], as well as capsid using only the capsid asymmetric unit [106]. As

investigate the relationship between pH and channel time has progressed, all-atom simulations of intact capsids

conductance [87,88] and internal water structure [89]. have become increasingly accessible. Notably, such work

Adamantane drug compounds (i.e. and has included characterization of the stress-response of the

rimantidine) are known to inhibit M2 channel activity southern bean mosaic virus (SBMV) capsid [109], valida-

[79]. Additional all-atom MD simulation work on M2 has tion of multi-scale modeling approaches based on the

Current Opinion in Virology 2018, 31:82–91 www.sciencedirect.com

All-atom simulations of viruses Hadden and Perilla 85



Sesbania mosaic virus (SeMV) capsid [110], derivation of guided MD simulations [118 ]. This landmark simula-

a complete atomic model for the rabbit hemorrhagic tion endeavor, encompassing 64 million atoms with sol-

disease virus (RHDV) capsid [111], and examination of vent, overcame the challenges of HIV-1 capsid size and

the interplay between solvent and ions and the porcine asymmetric architecture to generate the first chemically

circovirus type 2 (PCV2) capsid [112,113]. complete model. The results of the study revealed key

structural elements defining pentamer–hexamer and

The first all-atom simulation of a virus capsid to explore hexamer–hexamer interfaces within the capsid and pro-

the microsecond timescale was based on an empty model duced a much-needed platform to enable investigation of

of satellite tobacco necrosis virus (STNV), which encom- capsid function and targeted drug development. Notably,



passed 1.2 million atoms with solvent [114 ]. Like many the all-atom model has been used to study capsid inter-

non-enveloped plant viruses, STNV exhibits specific actions with the host cell factors cyclophilin A [119,120]

binding sites for divalent ions, likely calcium, and has and MxB [121], which regulate and restrict HIV-1 viral

been shown experimentally to undergo structural expan- infectivity, respectively.

sion unless these ions are bound. Simulations with and

without bound calcium confirmed that the former main- In a subsequent study, the empty HIV-1 capsid model

tain a smaller diameter and the latter swell. The results of was simulated for 1.2 ms, setting the current published

the work produced an atomistic model for the swollen record for the most substantial all-atom simulation ever



STNV capsid, which was not previously available, as well performed [122 ]. The simulation revealed that the

as an atomistic description of the dynamics underlying capsid translocates chloride across its surface at twice

the swelling process. the rate of sodium through ion-specific channels, which

may relate to the translocation of nucleotides during

The empty poliovirus capsid was studied over a timescale reverse transcription. Further, the capsid exhibited com-

of 200 ns, based on a system of 6 million atoms with plex dynamics, including collective motions that divide

solvent [115]. Simulation results revealed that the capsid the structure into two hemispheres and suggest a mecha-

functions as a semipermeable membrane structure, trans- nism for capsid uncoating, as well as oscillatory surface

locating water molecules, but not ions, in equilibrium waves at four different frequencies that may underlie

across its surface. Further, the solution pressure on the emergent properties of the capsid.

interior of the capsid was found to be negative, owing to

electrostatic interaction of solution electrolytes with the Immature particle lattices

charged capsid surface. The results of the work demon- such as HIV-1, human T-cell leukemia virus

strate that the capsid plays an essential role in maintaining type 1 (HTLV-1), and (RSV) initially

an environment conducive to the stable accommodation assemble as immature particles from Group-specific anti-

of its RNA genome and associated counterions. gen (Gag) polyproteins. During maturation, proteolytic

processing of Gag yields cleavage products that reassem-

The empty (HBV) capsid was simulated ble to produce mature virions [123]. All-atom simulations

for 1.1 ms based on a model of the complete assembly proved invaluable in determining the structures of the

domain, derived from crystallography and molecular immature HTLV-1 [124] and RSV [125] Gag lattices, as



modeling [116 ]. The simulation encompassed 6 million well as the HIV-1 capsid protein-SP1 peptide maturation



atoms with solvent and revealed that the capsid under- intermediate (CA-SP1) [126 ]. Notably, simulations

goes notable asymmetric distortion at equilibrium. Fur- revealed that the CA-SP1 six-helix bundle exists in a

ther, the capsid exhibited a fivefold preference to trans- dynamic helix-coil equilibrium, and that maturation-inhi-

locate sodium over chloride through its triangular pores, biting drugs and mutations act by stabilizing a helical



and induced the localization of sodium along its interior form of the bundle [126 ]. MD studies of CA-SP1 bound

surface. The results of the work implicate the triangular to inositol phosphates showed that these small molecules

pores as the extrusion site of the capsid’s carboxy-termi- facilitate the formation of the six-helix bundle and, thus,



nal domain tails, which contain cellular signals, and promote assembly of the Gag lattice [127 ]. Simulations

suggest a mechanism by which the capsid could modulate of the RSV lattice model were used to determine the

signal exposure based on tail phosphorylation. Simula- effects of charge distribution and mutations on Gag

tions have also been used to investigate the effects of assembly [128].



small-molecule drugs on HBV capsid morphology [107 ]

and to determine the structural basis for enhanced assem- Viral enzymes

bly and drug resistance in HBV capsid mutants [117]. Some viruses encode and package enzymes necessary to

carry out the processes that drive their infection cycles.

An all-atom structure of the human immunodeficiency All-atom MD simulations of the HIV-1 protease

virus type 1 (HIV-1) capsid (Figures 1b and 2 e) was (Figures 1b and 2 b), which is responsible for proteolytic

derived by combining results from cryo-EM and NMR cleavage of Gag prior to maturation, revealed the motion

experiments with computational modeling and data- of the enzyme’s active-site flaps, whose opening and

www.sciencedirect.com Current Opinion in Virology 2018, 31:82–91

86 Virus structure and expression

closing are considered essential to function [129–134]. bacteriophage MS2 [142,143]. The availability of more

Further, studies of the West Nile virus (WNV) NS3 complete structural information, along with improved

protease identified a conformational selection mechanism understanding of genome organization within capsids



used to identify its substrate [135]. Additionally, all-atom [144] and integrative modeling approaches [1 ] that

simulations were instrumental in elucidating the mecha- employ increasingly accurate protein and nucleic acid

nism by which non-nucleoside reverse transcription inhi- force fields [145,146], will support new simulation studies

bitors (NNRTIs), an important class of therapeutics of all-atom virions in the near future, particularly for non-

against HIV-1, interfere with the virus’ reverse transcri- enveloped viruses.

pase [136] (Figures 1b and 2 c), which synthesizes DNA

from the RNA template. All-atom simulations of enveloped viruses will face the

additional challenge of treating the envelope membrane

Toward all-atom simulations of complete lipid bilayer. Due to size, complex lipid compositions, and

virions the inclusion of membrane-embedded surface proteins,

As experimental methods, particularly cryo-EM and cryo- viral envelopes require careful consideration to model and

ET, continue to provide greater detail on virus composi- equilibrate [147]. Further, enclosing a virus capsid in a

tion and architecture, and as supercomputers and simula- fully atomistic envelope will significantly increase system

tion codes continue to offer greater performance for the size and simulation expense, particularly given the inclu-

study of large biological systems, the computational com- sion of solvent. Figure 2 emphasizes the dramatic size

munity becomes poised to study the dynamics of com- disparity between several protein-based components of

plete virions within physiological environments. None- HIV-1 that have been previously investigated with all-

theless, the paucity of accurate, full-length, and fully atom simulations and a model of the intact envelope

atomistic models for some major components of virus membrane (Figure 2f). The envelope necessarily dwarfs

structure presents a significant challenge to the derivation the HIV-1 capsid, the simulations of which currently

and investigation of all-atom virions. represent the most far-reaching accomplishment of

computational virology reported for structures of atomic

 

Importantly, the lack of accurate all-atom models for detail [118 ,122 ]. Following construction of a complete

encapsidated genome structures is the foremost limiting HIV-1 virion, it is clear that substantial supercomputing

factor for the construction of complete virions. While the power will be required to simulate such a colossal biolog-

work of Freddolino et al. on genome-containing STMV ical assembly at chemical resolution, especially over time-

(Figure 1c) represents the only all-atom simulation of a scales sufficient to allow meaningful study.



complete virion performed to date [108 ], the genome

structure employed was not based on the actual STMV

RNA sequence. Similarly, a complete pariacoto virus Figure 2

(PaV) virion derived using molecular modeling included

an RNA model based on an artificial RNA sequence [137]. (a) (e)

For both of these virions, limitations in determining

genome organization from crystal structures, which typi- (f)

cally rely on icosahedral averaging, led to inaccuracies in

(b)

the resulting RNA models [138,139]. Recently, Zeng et al.

used computational methods to generate the first all-atom

model for the complete structure of any virus using the

(c)

natural genome sequence, that of STMV containing

 realistic RNA [5 ].

(d)

Moving forward, modeling and simulation will continue

to play an essential role in the determination of all-atom

structures for genome-containing capsids, particularly

through approaches that integrate experimental data from (g)

techniques such as NMR and cryo-EM [140,141].

20 nm

Although simulation studies of the intact HBV capsid

have shown that the inherent flexibility of some virus

Current Opinion in Virology

structures may limit the ability to obtain true atomic (1–

˚ 

2 A) resolution with cryo-EM [116 ], advances in image

Graphical size comparison of viral protein components. (a) HIV-1

processing are enabling high-resolution asymmetric

viroporin (Vpu), transmembrane domain. (b) HIV-1 protease. (c) HIV-1

reconstructions capable of producing more authentic

reverse transcriptase (RT). (d) HIV-1 envelope protein (Env)

descriptions of encapsidated genome, such as those ectodomain. (e) HIV-1 capsid. HIV-1 envelope membrane. (g) STMV

obtained for the immature HBV particle and capsid.

Current Opinion in Virology 2018, 31:82–91 www.sciencedirect.com

All-atom simulations of viruses Hadden and Perilla 87

All-atom modeling of STMV with its encapsidated RNA genome, produ-

In the meantime, three viral envelopes have been inves-

cing the first realistic and chemically-complete structure of any virus.

tigated using coarse-grained simulation methods, includ-

6. Kasson P.M.: Receptor binding by influenza virus: using

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In addition to suitable numbers and distributions of

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for avian and human sialopentasaccharide receptor

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Insights into the human glycan receptor conformation of

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1918 pandemic hemagglutinin–glycan complexes derived

efforts. Additionally, experimental characterization of from nuclear magnetic resonance and molecular dynamics

studies. Biochemistry 2014, 53:4122-4135.

viral envelope lipidomes and improved software and force

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Acknowledgments

14. De Vries R.P., Peng W, Grant OC, Thompson AJ, Zhu X,

This work was supported by the National Institutes of Health grants

Bouwman KM, Torrents de la Pena AT, van Breemen M.J.,

P50GM082251 and P30GM110758-04 and the University of Delaware.

Ambepitiya Wickra-masinghe IN, de Haan C.A.M. et al.: Three

mutations switch H7N9 influenza to human-type receptor

specificity. PLoS Pathogens 2017, 13:e1006390.

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