Biology subjects

Voth, G. A.

Publications and source records attributed to Voth, G. A..

At least 37 records · Page 2Linked to original sources

bioRxiv2025

Mechanism for oil-phase separation by the lipid droplet assembly complex

Cells store metabolic energy as triglyceride (TG) oils in lipid droplets (LDs). LDs form de novo from the endoplasmic reticulum. How the lipid droplet assembly complex (LDAC), composed of seipin and LDAF11,2, catalyzes the organized formation of an oil phase in a membrane bilayer before spontaneous phase separation is triggered is unknown. Here, we reconstitute LD formation in vitro using purified LDAC and membranes containing physiologic levels of TG, demonstrating that the LDAC is both necessary and sufficient to catalyze oil-phase formation below the threshold of spontaneous phase separation. Structural studies of the LDAC reveal that LDAF1 forms a central ring within a seipin cage, creating a toroidal, membrane-spanning structure. Molecular dynamics simulations and biochemical assays show that this structure forms a selective chamber within the ER bilayer that limits phospholipids but allows TG to access a reaction compartment between the inner and outer rings of the LDAC. Within this compartment, TG interacts with LDAF1 and each other to form an oil phase to initiate LD formation. Thus, the LDAC acts as a protein catalyst for oil-phase separation in cells, revealing a fundamental mechanism for how cells resolve the biophysical challenge of storing oils within a hydrophilic environment in an organized manner.

cell biology↗

bioRxiv2025

Mechanistic Insights into Lenacapavir-Induced Off-Pathway HIV-1 Capsid Assembly

The HIV-1 capsid is a fullerene-like shell composed of hexamer and pentamer arrangements of the capsid (CA) proteins. The cone shape of the capsid is particularly important for packaging the viral genome and coordinating nuclear entry. Lenacapavir (LEN), a potent long-acting inhibitor, has been shown to disrupt capsid morphogenesis by binding at the FG-binding pocket located between neighboring CA subunits. Interestingly, inositol hexakisphosphate (IP6), a cellular polyanion, binds within the central pore of capsid pentamers and some hexamers while playing a key role in regulating the hexamer/pentamer switch. As LEN and IP6 interact with overlapping structural elements, they can compete to influence the capsid assembly pathway and outcomes. Using coarse-grained molecular simulations, we examined capsid assembly across varying IP6 and LEN conditions. Our results reveal a concentration-dependent shift in assembly outcomes: LEN accelerates hexamer assembly and reduces pentamer incorporation, leading to malformed, multilayered, or incomplete capsids. Simulations including a model for the viral ribonucleoprotein (RNP) complex further show that LEN-treated capsids frequently fail to encapsidate the RNA genome, indicating impaired maturation. Our calculations confirm that LEN impairs the formation of high-curvature CA lattice regions necessary for closure, supporting a model of off-pathway assembly as a mechanism of viral inhibition.

biophysics↗

bioRxiv2025

On the Curvature and Relaxation of Microtubule Plus-endTips

Microtubules are essential cytoskeletal components with a broad range of functions in which the structure and dynamics of their plus-end tips play critical roles. Existing mechanistic models explain the tips curving dynamics in different ways: the allosteric model suggests that GTP hydrolysis induces conformational changes in tubulin subunits that destabilize the lattice, leading to protofilament curving and depolymerization, while the lattice model posits that GTP hydrolysis directly destabilizes the microtubule lattice. However, the effect of GTP hydrolysis on the curving dynamics of microtubule tips remains incompletely understood. In this study, we employed a multiscale modeling approach, combining all-atom molecular dynamics simulations with Brownian dynamics simulations, to investigate the relaxation of microtubule plus-end tips into curved configurations. Our results show that both GDP- and GTP-bound tips exhibit an outward bending of protofilaments into curved, rams horn-like structures, characterized by a linear relationship between curvature and distance from the plus-end tip. These observations align with experimental cryo-ET images of microtubule plus-end tips in different nucleotide states. Collectively, our findings suggest that the outward bending of protofilaments at the plus-end tip is an intrinsic feature of microtubules, independent of the nucleotide state. SIGNIFICANCEUnderstanding how microtubules change shape is crucial for elucidating key cellular processes such as cell division and shape maintenance, which are fundamental to both physiological function and disease progression. This study supports the concept that the microtubule plus-end tip relaxation does not align with models that couple shape changes to GTP hydrolysis, at least for the topmost tubulin heterodimers. By interfacing bottom-up multiscale modeling -- using the longest reported atomistic molecular dynamics simulations of microtubule tips-- with existing cryo-ET data, it is shown that protofilament bending operates independently of nucleotide hydrolysis and likely depolymerization. These findings highlight the need for a new conceptual framework that separates GTP hydrolysis from microtubule tip flaring.

biophysics↗

bioRxiv2025

Lenacapavir-induced Lattice Hyperstabilization is Central to HIV-1 Capsid Failure at the Nuclear Pore Complex and in the Cytoplasm

Lenacapavir (LEN) is the first HIV-1 capsid inhibitor approved for clinical use. It inhibits multiple steps of the viral life cycle; however, the molecular details of the effect of LEN on capsid structure and the mechanistic steps of the inhibition are not understood. Recent studies show that intact cone-shaped capsids and capsids with LEN-induced breaks can dock at nuclear pore complexes (NPC), but only intact capsids enter the nucleus. In this work, we combined large-scale coarse-grained molecular dynamics simulations and live-cell imaging to investigate the stepwise mechanism of docking of LEN-treated capsids into the NPC. Capsids bound to substoichiometric concentrations of LEN can reach the NPC central channel. As the capsid advances to the nuclear end, lattice defects are formed at the pentamer-hexamer interface - primarily at the narrower end - leading to pentamer dissociation. Dissociation of pentamers is detrimental to capsid integrity, leading to both rupture of the narrow end and destabilization of the hexamer-hexamer interface. Structural analysis of LEN-capsid complexes in our simulations demonstrates heterogeneous hyperstabilization and loss of the essential pliability of the capsid protein lattice. Live-cell imaging of HIV-1 cores labeled with two different fluorescent markers showed that LEN-treated ruptured capsids were docked at the NPC but were not imported into the nucleus. We conclude that LEN contributes to the loss of capsid elasticity and integrity, inhibiting HIV-1 nuclear entry and replication. Our findings demonstrate that altering viral material properties can be an effective strategy for designing antiviral drugs that disrupt viral core nuclear entry.

biophysics↗

bioRxiv2025

The PXBAR-domain protein SNX9 selectively sequesters PI(3,4)P2 lipid and protects it from hydrolysis at the plasma membrane

Plasma membrane remodeling processes are tightly regulated by the spatiotemporal distribution and dynamic conversion of phosphoinositidyl lipids (PIPs). This regulation is controlled by the recruitment of proteins such as sorting nexin 9 (SNX9), a key mediator of late-stage endocytosis and macropinocytosis. Using live cell imaging, in vitro reconstitution assays, and molecular dynamics simulations, we investigated how SNX9 distinguishes between PI(3,4)P2 and PI(4,5)P2, and the physiological relevance of this selectivity. Our results revealed that during macropinocytic membrane ruffling, SNX9 is recruited in a spatiotemporally coordinated manner with PI(3,4)P2, but not with PI(4,5)P2. While SNX9 induces comparably weak mechanical remodeling on model membranes containing either PIP2 species, it exhibits a clear selective binding to PI(3,4)P2, mediated by a non-canonical interface. Through mutational analysis of key residues involved in this sequestration, we further demonstrated that SNX9 protects PI(3,4)P2 from hydrolysis. Together, these results reveal a previously unrecognized mechanism of SNX9-PIP2 lipid interaction that underscores SNX9s pivotal role in coordinating membrane remodeling processes. TeaserCurvature sensing BAR protein SNX9 selectively sequesters PI(3,4)P2 lipids, acting as a checkpoint in cell membrane remodeling.

biophysics↗

bioRxiv2024

Kinetic Implications of IP6 Anion Binding on the Molecular Switch of the HIV-1 Capsid Assembly

HIV-1 capsid proteins (CA) self-assemble into a fullerene-shaped capsid, enabling cellular transport and nuclear entry of the viral genome. A structural switch comprising the Thr-Val-Gly-Gly (TVGG) motif either assumes a disordered coil or a 310 helix conformation to regulate hexamer or pentamer assembly, respectively. The cellular polyanion inositol hexakisphosphate (IP6) binds to a positively charged pore of CA capsomers rich in arginine and lysine residues mediated by electrostatic interactions. Both IP6 binding and TVGG coil-to-helix transition are essential for pentamer formation. However, the connection between IP6 binding and TVGG conformational switch remains unclear. Using extensive atomistic simulations, we show that IP6 imparts structural order at the central ring, which results in multiple kinetically controlled events leading to the coil- to-helix conformational change of the TVGG motif. IP6 facilitates the helix-to-coil transition by allowing the formation of intermediate conformations. Our results identify the key kinetic role of IP6 in HIV-1 pentamer formation.

biophysics↗

bioRxiv2024

The C-terminus of the multi-drug efflux pump EmrE prevents proton leak by gating transport

The model multi-drug efflux pump from Escherichia coli, EmrE, can perform multiple types of transport leading to different biological outcomes, conferring resistance to some drug substrates and enhancing susceptibility to others. While transporters have traditionally been classified as antiporters, symporters, or uniporters, there is growing recognition that some transporters may exhibit mixed modalities. This raises new questions about the regulation and mechanisms of these transporters. Here we show that the C-terminal tail of EmrE acts as a secondary gate, preventing proton leak in the absence of drug. Substrate binding unlocks this gate, allowing transport to proceed. Truncation of the C-terminal tail ({Delta}107-EmrE) leads to altered pH regulation of alternating access, an important kinetic step in the transport cycle, as measured by NMR. {Delta}107-EmrE has increased proton leak in proteoliposome assays and bacteria expressing this mutant have reduced growth. MD simulations of {Delta}107-EmrE show formation of a water wire from the open face of the transporter to the primary binding site in the core, facilitating proton leak. In WT-EmrE, the C-terminal tail forms specific interactions that block formation of the water wire. Together these data strongly support the C-terminus of EmrE acting as a secondary gate that regulates access to the primary binding site in the core of the transporter.

biophysics↗

bioRxiv2024

Direct lipid interactions control SARS-CoV-2 M protein conformational dynamics and virus assembly

M is the most abundant structural membrane protein in coronaviruses and is essential for the formation of infectious virus particles. SARS-CoV-2 M adopts two conformations, Mshort and Mlong, and regulated transition between states is hypothesized to coordinate viral assembly and budding. However, the factors that regulate M conformation and roles for each state are unknown. Here, we discover a direct M-sphingolipid interaction that controls M conformational dynamics and virus assembly. We show M binds Golgi-enriched anionic lipids including ceramide-1-phosphate (C1P). Molecular dynamics simulations show C1P interaction promotes a long to short transition and energetically stabilizes Mshort. Cryo-EM structures show C1P specifically binds Mshort at a conserved site bridging transmembrane and cytoplasmic regions. Disrupting Mshort-C1P interaction alters M subcellular localization, reduces interaction with Spike and E, and impairs subsequent virus-like particle cell entry. Together, these results show endogenous signaling lipids regulate M structure and support a model in which Mshort is stabilized in the early endomembrane system to organize other structural proteins prior to viral budding.

microbiology↗

bioRxiv2024

Data-Driven Equation-Free Dynamics Applied to Many-Protein Complexes: The Microtubule Tip Relaxation

Microtubules (MTs) constitute the largest components of the eukaryotic cytoskeleton and play crucial roles in various cellular processes, including mitosis and intracellular transport. The property allowing MTs to cater to such diverse roles is attributed to dynamic instability, which is coupled to the hydrolysis of GTP (guanosine-5-triphosphate) to GDP (guanosine-5-diphosphate) within the {beta}-tubulin monomers. Understanding the equilibrium dynamics and the structural features of both GDP- and GTP-complexed MT tips, especially at an all-atom level, remains challenging for both experimental and computational methods because of their dynamic nature and the prohibitive computational demands of simulating large, many-protein systems. This study employs the "equation-free" multiscale computational method to accelerate the relaxation of all-atom simulations of MT tips toward their putative equilibrium conformation. Using large MT lattice systems (14 protofilaments x 8 heterodimers) comprising [~]21-38 million atoms, we applied this multiscale approach to leapfrog through time and nearly double the computational efficiency in realizing relaxed all-atom conformations of GDP- and GTP-complexed MT tips. Commencing from an initial 4 s unbiased all-atom simulation, we interleave coarse projective "equation-free" jumps with short bursts of all-atom molecular dynamics simulation to realize an additional effective simulation time of 1.875 s. Our 5.875 s of effective simulation trajectories for each system expose the subtle yet essential differences in the structures of MT tips as a function of whether {beta}-tubulin monomer is complexed with GDP or GTP, as well as the lateral interactions within the MT tip, offering a refined understanding of features underlying MT dynamic instability. The approach presents a robust and generalizable framework for future explorations of large biomolecular systems at atomic resolution. SIGNIFICANCEThe dynamic instability of microtubules (MTs) is essential for a plethora of biological functions, from cell division to intracellular transport. Despite their importance, current computational models often struggle to handle the large-scale, long-term dynamics of all-atom simulations. This computational study employs the "equation-free" method to accelerate relaxation of the all-atom structures of MT tips in both GDP and GTP states which nearly halves the computational demands of simulating very large biomolecular systems. Our findings expose subtle but crucial structural differences between GDP-bound and GTP-bound MTs, particularly in the number of protofilament clusters, and is consistent with a relatively stronger lateral interactions in GTP-bound MT tips.

biophysics↗

bioRxiv2024

Activation of the influenza B M2 proton channel (BM2)

Influenza B viruses have co-circulated during most seasonal flu epidemics and can cause significant human morbidity and mortality due to their rapid mutation, emerging drug resistance, and severe impact on vulnerable populations. The influenza B M2 proton channel (BM2) plays an essential role in viral replication, but the mechanisms behind its symmetric proton conductance and the involvement of a second histidine (His27) cluster remain unclear. Here we perform the membrane-enabled continuous constant-pH molecular dynamics simulations on wildtype BM2 and a key H27A mutant to explore its pH-dependent conformational switch. Simulations capture the activation as the first histidine (His19) protonates and reveal the transition at lower pH values compared to AM2 is a result of electrostatic repulsions between His19 and pre-protonated His27. Crucially, we provide an atomic-level understanding of the symmetric proton conduction by identifying pre-activating channel hydration in the C-terminal portion. This research advances our understanding of the function of BM2 function and lays the groundwork for further chemically reactive modeling of the explicit proton transport process as well as possible anti-flu drug design efforts.

biophysics↗

bioRxiv2024

Lipid Organization by the Caveolin-1 Complex

Caveolins are lipid-binding proteins that can organize membrane remodeling and oligomerize into the 8S-complex. The CAV1 8S-complex comprises a disk-like structure, about 15nm in diameter, with a central beta barrel. Further oligomerization of 8S-complexes remodels the membrane into caveolae vessels, with a dependence on cholesterol concentration. However, the molecular mechanisms behind membrane remodeling and cholesterol filtering are still not understood. Performing atomistic Molecular Dynamics simulations in combination with advanced sampling techniques, we describe how the CAV1-8S complex bends the membrane and accumulates cholesterol. Here, our simulations show an enhancing effect by the palmitoylations of CAV1, and we predict that the CAV1-8S complex can extract cholesterol molecules from the lipid bilayer and accommodate them in its beta barrel. Through backmapping to the all-atom level we also conclude that the Martini v2 coarse-grained forcefield overestimates membrane bending, as the atomistic simulations exhibit only very localized bending.

biophysics↗

bioRxiv2023

Divergent spike mutations impact the activation of the fusion core in Delta and Omicronvariants of SARS-CoV-2

SARS-CoV-2 infects host cells by binding the receptor-binding domain (RBD) of its spike protein to the receptor, ACE2. A subset of highly effective spike mutations plays critical roles in altering the conformational dynamics of spike protein. Here, we use molecular dynamics simulations to investigate how spike mutations affect the conformational dynamics of spike/ACE2 complex in the D614G, Delta (B.1.617.2) and Omicron (B.1.1.529) SARS-CoV-2 variants. We observe that the increased positive-charged mutations in the Omicron spike amplify its structural rigidity and reduce its structural flexibility. The mutations (P681R in Delta and P681H in Omicron) at the S1/S2 junction facilitate S1/S2 cleavage and aid the activation of the fusion core. We report that high structural flexibility in Delta lowers the barrier for the activation of the S2 core; however, high structural rigidity in Omicron enhances the barrier for the same. Our results also explain why Omicron requires the presence of a higher number of ACE2 to activate its fusion core than Delta.

biophysics↗

bioRxiv2023

Molecular dynamics simulations of HIV-1 matrix-membrane interactions at different stages of viral maturation

Although the structural rearrangement of the membrane-bound matrix (MA) protein trimers upon HIV-1 maturation has been reported, the consequences of MA maturation on the MA-lipid interactions are not well understood. Long-timescale molecular dynamics (MD) simulations of the MA multimeric assemblies of immature and mature virus particles with our realistic asymmetric membrane model have explored MA-lipid interactions and lateral organization of lipids around MA complexes. The number of stable MA-PS and MA-PIP2 interactions at the trimeric interface of the mature MA complex is observed to be greater compared to that of the immature MA complex. Our simulations identified an alternative PIP2 binding site in the immature MA complex where the multivalent headgroup of a PIP2 lipid with a greater negative charge binds to multiple basic amino acid residues such as, ARG3 residues of both the MA monomers at the trimeric interface, and HBR residues (LYS29, LYS31) of one of the MA monomers. Our enhanced sampling simulations have explored the conformational space of phospholipids at different binding sites of the trimeric interface of MA complexes. Unlike the immature MA complex, the 2 acyl tail of two PIP2 lipids at the trimeric interface of the mature MA complex is observed to sample stable binding pockets of MA consisting of helix4 residues. Together, our results provide molecular-level insights into the interactions of MA trimeric complexes with membrane and different lipid conformations at the specific binding sites of MA protein before and after viral maturation. SignificanceHIV-1 maturation facilitates the conversion of a newly formed immature virus particle to a mature infectious virion and initiates a new round of infection. The contributions of specific protein-lipid interactions in the HIV-1 assembly process are well recognized, however, the interactions of matrix protein lattice with the membrane before and after HIV-1 maturation are yet to be fully understood. Based on our simulated data, supported by prior experimental observations, the present study provides a molecular-level understanding of possible altered binding mode of PIP2 lipids after viral maturation. Identification of protein-lipid specific interactions, and lipid sorting data obtained from our long-time and large-scale atomistic MD simulations advance the understanding of the HIV-1 matrix and membrane maturation.

biophysics↗

bioRxiv2023

Cooperative Membrane Binding of HIV-1 Matrix Proteins

The HIV-1 assembly process begins with a newly synthesized Gag polyprotein being targeted to the inner leaflet of the plasma membrane of the infected cells to form immature viral particles. Gag-membrane interactions are mediated through the myristoylated(Myr) N-terminal matrix (MA) domain of Gag which eventually multimerize on the membrane to form trimers and higher-order oligomers. The study of the structure and dynamics of peripheral membrane proteins like MA has been challenging for both experimental and computational studies due to the complex dynamics of protein-membrane interactions. Although the roles of anionic phospholipids (PIP2, PS) and the Myr group in the membrane targeting and stable membrane binding of MA are now well-established, the cooperative interactions between MA monomers and MA-membrane still remain elusive. Our present study focuses on the membrane binding dynamics of a higher-order oligomeric structure of MA protein (a dimer of trimers), which has not been explored before. Employing time-lagged independent component analysis (tICA) to our microsecond-long trajectories, we investigate conformational changes of the matrix protein induced by membrane binding. Interestingly, the Myr switch of a MA monomer correlates with the conformational switch of adjacent monomers in the same trimer. Together, our findings suggest that MA trimerization facilitates Myr insertion, but MA trimer-trimer interactions in the lattice of immature HIV-1 particles can hinder the same. Additionally, local lipid density patterns of different lipid species provide a signature of the initial stage of lipid-domain formation upon membrane binding of the protein complex. TOC O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=104 SRC="FIGDIR/small/559012v1_ufig1.gif" ALT="Figure 1"> View larger version (51K): org.highwire.dtl.DTLVardef@c4380corg.highwire.dtl.DTLVardef@3bbc05org.highwire.dtl.DTLVardef@1b50f42org.highwire.dtl.DTLVardef@1fccd7f_HPS_FORMAT_FIGEXP M_FIG C_FIG

biophysics↗

bioRxiv2023

The SARS-CoV-2 nucleoprotein associates with anionic lipid membranes

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a lipid-enveloped virus that acquires its lipid bilayer from the host cell it infects. SARS-CoV-2 can spread from cell to cell or from patient to patient by undergoing assembly and budding to form new virions. The assembly and budding of SARS-CoV-2 is mediated by several structural proteins known as envelope (E), membrane (M), nucleoprotein (N) and spike (S), which can form virus-like particles (VLPs) when co-expressed in mammalian cells. Assembly and budding of SARS-CoV-2 from the host ER-Golgi intermediate compartment is a critical step in the virus acquiring its lipid bilayer. To date, little information is available on how SARS-CoV-2 assembles and forms new viral particles from host membranes. In this study, we find the N protein can strongly associate with anionic lipids including phosphoinositides and phosphatidylserine. Moreover, lipid binding is shown to occur in the N protein C-terminal domain, which is supported by extensive in silico analysis. Anionic lipid binding occurs for both the free and N oligomeric forms suggesting N can associate with membranes in the nucleocapsid form. Herein we present a lipid-dependent model based on in vitro, cellular and in silico data for the recruitment of N to M assembly sites in the lifecycle of SARS-CoV-2.

microbiology↗

bioRxiv2023

Conformational transitions of the HIV-1 Gag polyprotein upon multimerization and gRNA binding

During the HIV-1 assembly process, the Gag polyprotein multimerizes at the producer cell plasma membrane, resulting in the formation of spherical immature virus particles. Gag-gRNA interactions play a crucial role in the multimerization process, which is yet to be fully understood. We have performed large-scale all-atom molecular dynamics simulations of membrane-bound full-length Gag dimer, hexamer, and 18-mer. The inter-domain dynamic correlation of Gag, quantified by the heterogeneous elastic network model (hENM) applied to the simulated trajectories, is observed to be altered by implicit gRNA binding, as well as the multimerization state of the Gag. The lateral dynamics of our simulated membrane-bound Gag proteins, with and without gRNA binding, agree with prior experimental data and help to validate our simulation models and methods. The gRNA binding is observed to impact mainly the SP1 domain of the 18-mer and the MA-CA linker domain of the hexamer. In the absence of gRNA binding, the independent dynamical motion of the NC domain results in a collapsed state of the dimeric Gag. Unlike stable SP1 helices in the six-helix bundle, without IP6 binding, the SP1 domain undergoes a spontaneous helix-to-coil transition in the dimeric Gag. Together, our findings reveal conformational switches of Gag at different stages of the multimerization process and predict that the gRNA binding reinforces an efficient binding surface of Gag for multimerization, as well as regulates the dynamic organization of the local membrane region itself. SignificanceGag(Pr55Gag) polyprotein orchestrates many essential events in HIV-1 assembly, including packaging of the genomic RNA (gRNA) in the immature virion. Although various experimental techniques, such as cryo-ET, X-ray, and NMR, have revealed structural properties of individual domains in the immature Gag clusters, structural and biophysical characterization of a full-length Gag molecule remains a challenge for existing experimental techniques. Using atomistic molecular dynamics simulations of the different model systems of Gag polyprotein, we present here a detailed structural characterization of Gag molecules in different multimerization states and interrogate the synergy between Gag-Gag, Gag-membrane, and Gag-gRNA interactions during the viral assembly process.

biophysics↗

bioRxiv2023

HIV-1 Capsid Shape, Orientation, and Entropic Elasticity Regulate Translocationinto the Nuclear Pore Complex

Nuclear import of the viral capsid is a critical step in the HIV-1 life cycle that serve to transport and release genomic material into the nucleus. Nuclear Pore Complex (NPC) allows passage of intact capsid, though mechanistic details of the process remain to be fully understood. Here we investigate the factors regulating HIV-1 capsid translocation into the NPC central channel using coarse-grained molecular dynamics simulations. We find that successful translocation is contingent on the compatibility of the capsid morphology and channel dimension and the proper orientation of the capsid approach to the channel. The central channel dynamically expands to allow capsid passage, demonstrating the pleomorphic nature of the channel necessary for transporting large cargoes. Structural analysis shows that stress induced by the central channel confinement and uncondensed internal genomic material generates correlated striated patterns of lattice disorder across the viral capsid surface which is an indicator of its lattice "elasticity". Our results suggest that the "elasticity" can aid the capsid to adapt to the stress and remain structurally intact during translocation. TeaserComputer Simulations identify capsid elasticity as a key factor for successful nuclear entry.

biophysics↗

bioRxiv2023

Mechanism of Phosphate Release from Actin Filaments

After ATP-actin monomers assemble filaments, the ATPs {gamma}-phosphate is hydrolyzed within seconds and dissociates over minutes. We used all-atom molecular dynamics simulations to sample the release of phosphate from filaments and study residues that gate release. Dissociation of phosphate from Mg2+ is rate limiting and associated with an energy barrier of 20 kcal/mol, consistent with experimental rates of phosphate release. Phosphate then diffuses in an internal cavity toward a gate formed by R177 suggested in prior computational studies and cryo-EM structures. The gate is closed when R177 hydrogen bonds with N111 and is open when R177 forms a salt bridge with D179. Most of the time interactions of R177 with other residues occludes the phosphate release pathway. Machine learning analysis reveals that the occluding interactions fluctuate rapidly, underscoring the secondary role of backdoor gate opening in Pi release, in contrast with the previous hypothesis that gate opening is the primary event. Significance StatementThe protein actin assembles into filaments that participate in muscle contraction and cellular movements. An ATP bound to the actin monomer is hydrolyzed rapidly during filament assembly, but the {gamma}-phosphate dissociates slowly from the filament. We identified phosphate dissociation from Mg2+ as the rate-limiting step in phosphate release from actin based on an energy barrier that aligns with the experimentally determined release rate. The release of phosphate from the protein requires opening a gate in the actin molecule formed by the interaction between sidechains of arginine 177 and asparagine 111. Surprisingly, simulations revealed other interactions of the sidechain of arginine 177 that occlude the release pathway most of the time but have not been observed in low-temperature cryo-EM structures.

biophysics↗