Multiscale Molecular Dynamics Simulations Research Articles

One molecule to couple them all: Toward realistic numbers of molecules in multiscale molecular dynamics simulations of exciton-polaritons.

Collective strong coupling of many molecules to the confined light modes of an optical resonator can influence the photochemistry of these molecules, but the origin of this effect is not yet fully understood. To provide atomistic insights, several approaches have been developed based on quantum chemistry or molecular dynamics methods. However, most of these methods rely on coupling a few molecules (or sometimes only one) to a single cavity mode. To reach the strong coupling regime with such a small number of molecules, much larger vacuum field strengths are employed than in experiments. To keep the vacuum field realistic and avoid potential artefacts, the number of coupled molecules should be significantly increased instead, but that is not always possible due to restrictions on computational hardware and software. To overcome this barrier and model the dynamics of an arbitrarily large ensemble of molecules coupled to realistic cavity fields in atomistic molecular dynamics simulations, we propose to coarse-grain subsets of molecules into one or more effective supermolecules with an enhanced dipole moment and concerted dynamics. To verify the validity of the proposed multiscale model, we performed simulations in which we investigated how the number of molecules that are coupled to the cavity affects excited-state intra-molecular proton transfer, polariton relaxation, and exciton transport.

Lys716 in the transmembrane domain of yeast mitofusin Fzo1 modulates anchoring and fusion

Outer mitochondrial membrane fusion, a vital cellular process, is mediated by mitofusins. However, the underlying molecular mechanism remains elusive. We have performed extensive multiscale molecular dynamics simulations to predict a model of the transmembrane (TM) domain of the yeast mitofusin Fzo1. Coarse-grained simulations of the two TM domain helices, TM1 and TM2, reveal a stable interface, which is controlled by the charge status of residue Lys716. Atomistic replica-exchange simulations further tune our model, which is confirmed by a remarkable agreement with an independent AlphaFold2 (AF2) prediction of Fzo1 in complex with its fusion partner Ugo1. Furthermore, the presence of the TM domain destabilizes the membrane, even more if Lys716 is charged, which can be an asset for initiating fusion. The functional role of Lys716 was confirmed with yeast experiments, which show that mutating Lys716 to a hydrophobic residue prevents mitochondrial fusion.

Long-range charge transfer in respiratory complex I: insights from multi-scale molecular dynamics simulations

Long-range charge transfer in respiratory complex I: insights from multi-scale molecular dynamics simulations

Insertion and Anchoring of HIV-1 Fusion Peptide into Complex Membrane Mimicking Human T-cell.

A fundamental understanding of how HIV-1 envelope (Env) protein facilitates fusion is still lacking. The HIV-1 fusion peptide, consisting of 15 to 22 residues, is the N-terminus of the gp41 subunit of the Env protein. Further, this peptide, a promising vaccine candidate, initiates viral entry into target cells by inserting and anchoring into human immune cells. The influence of membrane lipid reorganization and the conformational changes of the fusion peptide during the membrane insertion and anchoring processes, which can significantly affect HIV-1 cell entry, remains largely unexplored due to the limitations of experimental measurements. In this work, we investigate the insertion of the fusion peptide into an immune cell membrane mimic through multiscale molecular dynamics simulations. We mimic the native T-cell by constructing a 9-lipid asymmetric membrane, along with geometrical restraints accounting for insertion in the context of gp41. To account for the slow timescale of lipid mixing while enabling conformational changes, we implement a protocol to go back and forth between atomistic and coarse-grained simulations. Our study provides a molecular understanding of the interactions between the HIV-1 fusion peptide and the T-cell membrane, highlighting the importance of conformational flexibility of fusion peptides and local lipid reorganization in stabilizing the anchoring of gp41 into the targeted host membrane during the early events of HIV-1 cell entry. Importantly, we identify a motif within the fusion peptide critical for fusion that can be further manipulated in future immunological studies.

Impact of solvation on the photoisomerisation dynamics of a photon-only rotary molecular motor

The optimization of the quantum efficiency of single-molecule light-driven rotary motors typically relies on chemical modifications. While, in isolated conditions, computational methods have been frequently used to design more efficient motors, the role played by the solvent environment has not been satisfactorily investigated. In this study, we used multiscale nonadiabatic molecular dynamics simulations of the working cycle of a 2-stroke photon-only molecular rotary motor. The results, which display dynamics consistent with the available transient spectroscopy measurements, predict a considerable decrease in the isomerisation quantum efficiency in methanol solution with respect to the gas phase. The origin of such a decrease is traced back to the ability of the motor to establish hydrogen bonds with solvent molecules. The analysis suggests that a modified motor with a reduced ability to form hydrogen bonds will display increased quantum efficiency, therefore extending the set of engineering rules available for designing light-driven rotary motors.

GENESIS 2.1: High-Performance Molecular Dynamics Software for Enhanced Sampling and Free-Energy Calculations for Atomistic, Coarse-Grained, and Quantum Mechanics/Molecular Mechanics Models.

GENeralized-Ensemble SImulation System (GENESIS) is a molecular dynamics (MD) software developed to simulate the conformational dynamics of a single biomolecule, as well as molecular interactions in large biomolecular assemblies and between multiple biomolecules in cellular environments. To achieve the latter purpose, the earlier versions of GENESIS emphasized high performance in atomistic MD simulations on massively parallel supercomputers, with or without graphics processing units (GPUs). Here, we implemented multiscale MD simulations that include atomistic, coarse-grained, and hybrid quantum mechanics/molecular mechanics (QM/MM) calculations. They demonstrate high performance and are integrated with enhanced conformational sampling algorithms and free-energy calculations without using external programs except for the QM programs. In this article, we review new functions, molecular models, and other essential features in GENESIS version 2.1 and discuss ongoing developments for future releases.

Unraveling the Molecular Dance: Insights into TREM2/DAP12 Complex Formation in Alzheimer's Disease through Molecular Dynamics Simulations.

Alzheimer's disease (AD) is a widespread neurodegenerative condition affecting millions globally. Recent research has implicated variants of the triggering receptor expressed in myeloid cells 2 (TREM2) as risk factors for AD. TREM2, an immunomodulatory receptor on microglial surfaces, plays a pivotal role in regulating microglial activation by association with DNAX-activation protein 12 (DAP12). Despite its significance, the mechanism underlying the formation of the complex between the transmembrane domains (TMDs) of TREM2 and DAP12 remains unclear. This study employs multiscale molecular dynamics (MD) simulations to investigate three TMD complex models, including two derived from experiments and one generated by AlphaFold2. Conducted within a lipid membrane consisting of an 80:20 mixture of phosphatidylcholine (POPC) and cholesterol, our analysis reveals hydrogen-bonding interactions between K26 of TREM2 and D16 of DAP12 in all three models, consistent with previous experimental findings. Our results elucidate the different spatial conformations observed in the models and offer insights into the structure of the TREM2/DAP12 TMD complex. Furthermore, we elucidate the role of charged residues in the assembly structure of the complex within the lipid membrane. These findings enhance our understanding of the molecular mechanism governing TREM2/DAP12 complex formation, providing a foundation for designing novel therapeutic strategies to address AD and other neurodegenerative diseases.

State-dependent binding of cholesterol and an anionic lipid to the muscle-type Torpedo nicotinic acetylcholine receptor

The ability of the Torpedo nicotinic acetylcholine receptor (nAChR) to undergo agonist-induced conformational transitions requires the presence of cholesterol and/or anionic lipids. Here we use recently solved structures along with multiscale molecular dynamics simulations to examine lipid binding to the nAChR in bilayers that have defined effects on nAChR function. We examine how phosphatidic acid and cholesterol, lipids that support conformational transitions, individually compete for binding with phosphatidylcholine, a lipid that does not. We also examine how the two lipids work synergistically to stabilize an agonist-responsive nAChR. We identify rapidly exchanging lipid binding sites, including both phospholipid sites with a high affinity for phosphatidic acid and promiscuous cholesterol binding sites in the grooves between adjacent transmembrane α-helices. A high affinity cholesterol site is confirmed in the inner leaflet framed by a key tryptophan residue on the MX α-helix. Our data provide insight into the dynamic nature of lipid-nAChR interactions and set the stage for a detailed understanding of the mechanisms by which lipids facilitate nAChR function at the neuromuscular junction.

A robust computational quest: Discovering potential hits to improve the treatment of pyrazinamide-resistant Mycobacterium tuberculosis.

The rise of pyrazinamide (PZA)-resistant strains of Mycobacterium tuberculosis (MTB) poses a major challenge to conventional tuberculosis (TB) treatments. PZA, a cornerstone of TB therapy, must be activated by the mycobacterial enzyme pyrazinamidase (PZase) to convert its active form, pyrazinoic acid, which targets the ribosomal protein S1. Resistance, often associated with mutations in the RpsA protein, complicates treatment and highlights a critical gap in the understanding of structural dynamics and mechanisms of resistance, particularly in the context of the G97D mutation. This study utilizes a novel integration of computational techniques, including multiscale biomolecular and molecular dynamics simulations, physicochemical and medicinal chemistry predictions, quantum computations and virtual screening from the ZINC and Chembridge databases, to elucidate the resistance mechanism and identify lead compounds that have the potential to improve treatment outcomes for PZA-resistant MTB, namely ZINC15913786, ZINC20735155, Chem10269711, Chem10279789 and Chem10295790. These computational methods offer a cost-effective, rapid alternative to traditional drug trials by bypassing the need for organic subjects while providing highly accurate insight into the binding sites and efficacy of new drug candidates. The need for rapid and appropriate drug development emphasizes the need for robust computational analysis to justify further validation through invitro and invivo experiments.

Mechanistic Insight into the Mechanical Unfolding of the Integral Membrane Diacylglycerol Kinase.

The essential forces stabilizing membrane proteins and governing their folding and unfolding are difficult to decipher. Single-molecule atomic force spectroscopy mechanically unfolds individual membrane proteins and quantifies their dynamics and energetics. However, it remains challenging to structurally assign unfolding intermediates precisely and to deduce dominant interactions between specific residues that facilitate either the localized stabilization of these intermediates or the global assembly of membrane proteins. Here, we performed force spectroscopy experiments and multiscale molecular dynamics simulations to study the unfolding pathway of diacylglycerol kinase (DGK), a small trimeric multispan transmembrane enzyme. The remarkable agreement between experiments and simulations allowed precise structural assignment and interaction analysis of unfolding intermediates, bypassing existing limitations on structural mapping, and thus provided mechanistic explanations for the formation of these states. DGK unfolding was found to proceed with structural segments varying in size that do not correlate with its secondary structure. We identified intermolecular side-chain packing interactions as one of the major contributions to the stability of unfolding intermediates. Mutagenesis creating packing defects induced a dramatic decrease in the mechano-stability of corresponding intermediates and also in the thermo-stability of DGK trimer, in good agreement with predictions from simulations. Hence, the molecular determinants of the mechano- and thermo-stability of a membrane protein can be identified at residue resolution. The accurate structural assignment established and microscopic mechanism revealed in this work may substantially expand the scope of single-molecule studies of membrane proteins.

A binding site for phosphoinositides described by multiscale simulations explains their modulation of voltage-gated sodium channels

Voltage-gated sodium channels (Naᵥ) are membrane proteins which open to facilitate the inward flux of sodium ions into excitable cells. In response to stimuli, Naᵥ channels transition from the resting, closed state to an open, conductive state, before rapidly inactivating. Dysregulation of this functional cycle due to mutations causes diseases including epilepsy, pain conditions, and cardiac disorders, making Naᵥ channels a significant pharmacological target. Phosphoinositides are important lipid cofactors for ion channel function. The phosphoinositide PI(4,5)P2 decreases Naᵥ1.4 activity by increasing the difficulty of channel opening, accelerating fast inactivation and slowing recovery from fast inactivation. Using multiscale molecular dynamics simulations, we show that PI(4,5)P2 binds stably to inactivated Naᵥ at a conserved site within the DIV S4–S5 linker, which couples the voltage-sensing domain (VSD) to the pore. As the Naᵥ C-terminal domain is proposed to also bind here during recovery from inactivation, we hypothesize that PI(4,5)P2 prolongs inactivation by competitively binding to this site. In atomistic simulations, PI(4,5)P2 reduces the mobility of both the DIV S4–S5 linker and the DIII–IV linker, responsible for fast inactivation, slowing the conformational changes required for the channel to recover to the resting state. We further show that in a resting state Naᵥ model, phosphoinositides bind to VSD gating charges, which may anchor them and impede VSD activation. Our results provide a mechanism by which phosphoinositides alter the voltage dependence of activation and the rate of recovery from inactivation, an important step for the development of novel therapies to treat Naᵥ-related diseases.

A binding site for phosphoinositides described by multiscale simulations explains their modulation of voltage-gated sodium channels.

Voltage-gated sodium channels (Naᵥ) are membrane proteins which open to facilitate the inward flux of sodium ions into excitable cells. In response to stimuli, Naᵥ channels transition from the resting, closed state to an open, conductive state, before rapidly inactivating. Dysregulation of this functional cycle due to mutations causes diseases including epilepsy, pain conditions, and cardiac disorders, making Naᵥ channels a significant pharmacological target. Phosphoinositides are important lipid cofactors for ion channel function. The phosphoinositide PI(4,5)P2 decreases Naᵥ1.4 activity by increasing the difficulty of channel opening, accelerating fast inactivation and slowing recovery from fast inactivation. Using multiscale molecular dynamics simulations, we show that PI(4,5)P2 binds stably to inactivated Naᵥ at a conserved site within the DIV S4-S5 linker, which couples the voltage-sensing domain (VSD) to the pore. As the Naᵥ C-terminal domain is proposed to also bind here during recovery from inactivation, we hypothesize that PI(4,5)P2 prolongs inactivation by competitively binding to this site. In atomistic simulations, PI(4,5)P2 reduces the mobility of both the DIV S4-S5 linker and the DIII-IV linker, responsible for fast inactivation, slowing the conformational changes required for the channel to recover to the resting state. We further show that in a resting state Naᵥ model, phosphoinositides bind to VSD gating charges, which may anchor them and impede VSD activation. Our results provide a mechanism by which phosphoinositides alter the voltage dependence of activation and the rate of recovery from inactivation, an important step for the development of novel therapies to treat Naᵥ-related diseases.

Understanding Self-Assembly and Molecular Packing in Methylcellulose Aqueous Solutions Using Multiscale Modeling and Simulations.

We present a multiscale molecular dynamics (MD) simulation study on self-assembly in methylcellulose (MC) aqueous solutions. First, using MD simulations with a new coarse-grained (CG) model of MC chains in implicit water, we establish how the MC chains self-assemble to form fibrils and fibrillar networks and elucidate the MC chains' packing within the assembled fibrils. The CG model for MC is extended from a previously developed model for unsubstituted cellulose and captures the directionality of H-bonding interactions between the -OH groups. The choice and placement of the CG beads within each monomer facilitates explicit modeling of the exact degree and position of methoxy substitutions in the monomers along the MC chain. CG MD simulations show that with increasing hydrophobic effect and/or increasing H-bonding strength, the commercial MC chains (with degree of methoxy substitution, DS, ∼1.8) assemble from a random dispersed configuration into fibrils. The assembled fibrils exhibit consistent fibril diameters regardless of the molecular weight and concentration of MC chains, in agreement with past experiments. Most MC chains' axes are aligned with the fibril axis, and some MC chains exhibit twisted conformations in the fibril. To understand the molecular driving force for the twist, we conduct atomistic simulations of MC chains preassembled in fibrils (without any chain twists) in explicit water at 300 and 348 K. These atomistic simulations also show that at DS = 1.8, MC chains adopt twisted conformations, with these twists being more prominent at higher temperatures, likely as a result of shielding of hydrophobic methyl groups from water. For MC chains with varying DS, at 348 K, atomistic simulations show a nonmonotonic effect of DS on water-monomer contacts. For 0.0 < DS < 0.6, the MC monomers have more water contacts than at DS = 0.0 or DS > 0.6, suggesting that with few methoxy substitutions, the MC chains are effectively hydrophilic, letting the water molecules diffuse into the fibril to participate in H-bonds with the MC chains' remaining -OH groups. At DS > 0.6, the MC monomers become increasingly hydrophobic, as seen by decreasing water contacts around each monomer. We conclude based on the atomistic observations that MC chains with lower degrees of substitutions (DS ≤ 0.6) should exhibit solubility in water over broader temperature ranges than DS ∼ 1.8 chains.

Lipid Landscapes: Vibrational Spectroscopy for Decoding Membrane Complexity.

Cell membranes are incredibly complex environments containing hundreds of components. Despite substantial advances in the past decade, fundamental questions related to lipid-lipid interactions and heterogeneity persist. This review explores the complexity of lipid membranes, showcasing recent advances in vibrational spectroscopy to characterize the structure, dynamics, and interactions at the membrane interface. We include an overview of modern techniques such as surface-enhanced infrared spectroscopy as a steady-state technique with single-bilayer sensitivity, two-dimensional sum-frequency generation spectroscopy, and two-dimensional infrared spectroscopy to measure time-evolving structures and dynamics with femtosecond time resolution. Furthermore, we discuss the potential of multiscale molecular dynamics (MD) simulations, focusing on recently developed simulation algorithms, which have emerged as a powerful approach to interpret complex spectra. We highlight the ongoing challenges in studying heterogeneous environments in multicomponent membranes via current vibrational spectroscopic techniques and MD simulations. Overall, this review provides an up-to-date comprehensive overview of the powerful combination of vibrational spectroscopy and simulations, which has great potential to illuminate lipid-lipid, lipid-protein, and lipid-water interactions in the intricate conformational landscape of cell membranes.

Reconciling ASPP-p53 binding mode discrepancies through an ensemble binding framework that bridges crystallography and NMR data.

ASPP2 and iASPP bind to p53 through their conserved ANK-SH3 domains to respectively promote and inhibit p53-dependent cell apoptosis. While crystallography has indicated that these two proteins employ distinct surfaces of their ANK-SH3 domains to bind to p53, solution NMR data has suggested similar surfaces. In this study, we employed multi-scale molecular dynamics (MD) simulations combined with free energy calculations to reconcile the discrepancy in the binding modes. We demonstrated that the binding mode based solely on a single crystal structure does not enable iASPP's RT loop to engage with p53's C-terminal linker-a verified interaction. Instead, an ensemble of simulated iASPP-p53 complexes facilitates this interaction. We showed that the ensemble-average inter-protein contacting residues and NMR-detected interfacial residues qualitatively overlap on ASPP proteins, and the ensemble-average binding free energies better match experimental KD values compared to single crystallgarphy-determined binding mode. For iASPP, the sampled ensemble complexes can be grouped into two classes, resembling the binding modes determined by crystallography and solution NMR. We thus propose that crystal packing shifts the equilibrium of binding modes towards the crystallography-determined one. Lastly, we showed that the ensemble binding complexes are sensitive to p53's intrinsically disordered regions (IDRs), attesting to experimental observations that these IDRs contribute to biological functions. Our results provide a dynamic and ensemble perspective for scrutinizing these important cancer-related protein-protein interactions (PPIs).

Multiscale Molecular Dynamics Simulations of an Active Self-Assembling Material.

Inspired by the adaptability observed in biological materials, self-assembly processes have attracted significant interest for their potential to yield novel materials with unique properties. However, experimental methods have often fallen short in capturing the molecular details of the assembly process. In this study, we employ a multiscale molecular dynamics simulation approach, complemented by NMR quantification, to investigate the mechanism of self-assembly in a redox-fueled bioinspired system. Contrary to conventional assumptions, we have uncovered a significant role played by the monomer precursor in the assembly process, with its presence varying with concentration and the extent of conversion of the monomer to the dimer. Experimental confirmation through NMR quantification underscores the concentration-dependent incorporation of monomers into the fibrous structures. Furthermore, our simulations also shed light on the diverse intermolecular interactions, including T-shaped and parallel π stacking, as well as hydrogen bonds, in stabilizing the aggregates. Overall, the open conformation of the dimer is preferred within these aggregates. However, inside the aggregates, the distribution of conformations shifts slightly to the closed conformation compared to on the surface. These findings contribute to the growing field of bioinspired materials science by providing valuable mechanistic and structural insights to guide the design and development of self-assembling materials with biomimetic functionalities.

The role of cryptic ancestral symmetry in histone folding mechanisms across Eukarya and Archaea.

Histones compact and store DNA in both Eukarya and Archaea, forming heterodimers in Eukarya and homodimers in Archaea. Despite this, the folding mechanism of histones across species remains unclear. Our study addresses this gap by investigating 11 types of histone and histone-like proteins across humans, Drosophila, and Archaea through multiscale molecular dynamics (MD) simulations, complemented by NMR and circular dichroism experiments. We confirm and elaborate on the widely applied "folding upon binding" mechanism of histone dimeric proteins and report a new alternative conformation, namely, the inverted non-native dimer, which may be a thermodynamically metastable configuration. Protein sequence analysis indicated that the inverted conformation arises from the hidden ancestral head-tail sequence symmetry underlying all histone proteins, which is congruent with the previously proposed histone evolution hypotheses. Finally, to explore the potential formations of homodimers in Eukarya, we utilized MD-based AWSEM and AI-based AlphaFold-Multimer models to predict their structures and conducted extensive all-atom MD simulations to examine their respective structural stabilities. Our results suggest that eukaryotic histones may also form stable homodimers, whereas their disordered tails bring significant structural asymmetry and tip the balance towards the formation of commonly observed heterotypic dimers.

Multiscale simulations reveal the driving forces of p53C phase separation accelerated by oncogenic mutations.

Liquid-Liquid phase separation (LLPS) of p53 to form liquid condensates has been implicated in cellular functions and dysfunctions. The p53 condensates may serve as amyloid fibril precursors to initiate p53 aggregation, which is associated with oncogenic gain-of-function and various human cancers. M237I and R249S mutations located in p53 core domain (p53C) have been detected respectively in glioblastomas and hepatocellular carcinoma. Interestingly, these p53C mutants can also undergo LLPS and liquid-to-solid phase transition, which are faster than wild type p53C. However, the underlying molecular basis governing the accelerated LLPS and liquid-to-solid transition of p53C remain poorly understood. Herein, we explore the M237I/R249S mutation-induced structural alterations and phase separation behavior of p53C by employing multiscale molecular dynamics simulations. All-atom simulations revealed conformational disruptions in the zinc-binding domain of the M237I mutant and in both loop3 and zinc-binding domain of the R249S mutant. The two mutations enhance hydrophobic exposure of those regions and attenuate intramolecular interactions, which may hasten the LLPS and aggregation of p53C. Martini 3 coarse-grained simulations demonstrated spontaneous phase separation of p53C and accelerated effects of M237I/R249S mutations on the phase separation of p53C. Importantly, we find that the regions with enhanced intermolecular interactions observed in coarse-grained simulations coincide with the disrupted regions with weakened intramolecular interactions observed in all-atom simulations, indicating that M237I/R249S mutation-induced local structural disruptions expedite the LLPS of p53C. This study unveils the molecular mechanisms underlying the two cancer-associated mutation-accelerated LLPS and aggregation of p53C, providing avenues for anticancer therapy by targeting the phase separation process.

Multiscale Molecular Dynamics Simulations of the Homodimer Accessory Protein ORF7b of SARS-CoV-2.

The SARS-CoV-2 ORF7b protein has drawn attention for its potential role in viral pathogenesis, but its structural details and lateral membrane associations remain elusive. In this study, we conducted multiscale molecular dynamics simulations to provide detailed molecular insights of the protein's dimerization, which is crucial for unraveling its structural model of protein-protein interface important to regulating cellular immune response. To gain a deeper understanding of homodimer configurations, we employed a machine learning algorithm for structural-based clustering. Clusters were categorized into three distinct groups for both parallel and antiparallel orientations, highlighting the influence of the initial monomer conformation on dimer configurations. Analysis of hydrogen bonding and π-π and π-cation stacking interactions within clusters revealed variations in interactions between clusters. In parallel dimers, weak stacking interactions in the transmembrane (TM) region were observed. In contrast, antiparallel dimers exhibited strong hydrogen bonding and stacking interactions contributing to tight dimeric packing, both within and outside the TM domain. Overall, our study provides a comprehensive view of the structural dynamics of ORF7b homodimerization in both parallel and antiparallel orientations. These findings shed light on the molecular interactions involved in ORF7b dimerization, which are crucial for understanding its potential roles in SARS-CoV-2 pathogenesis. This knowledge could inform future research and therapeutic strategies targeting this viral protein.

LipIDens: simulation assisted interpretation of lipid densities in cryo-EM structures of membrane proteins

Cryo-electron microscopy (cryo-EM) enables the determination of membrane protein structures in native-like environments. Characterising how membrane proteins interact with the surrounding membrane lipid environment is assisted by resolution of lipid-like densities visible in cryo-EM maps. Nevertheless, establishing the molecular identity of putative lipid and/or detergent densities remains challenging. Here we present LipIDens, a pipeline for molecular dynamics (MD) simulation-assisted interpretation of lipid and lipid-like densities in cryo-EM structures. The pipeline integrates the implementation and analysis of multi-scale MD simulations for identification, ranking and refinement of lipid binding poses which superpose onto cryo-EM map densities. Thus, LipIDens enables direct integration of experimental and computational structural approaches to facilitate the interpretation of lipid-like cryo-EM densities and to reveal the molecular identities of protein-lipid interactions within a bilayer environment. We demonstrate this by application of our open-source LipIDens code to ten diverse membrane protein structures which exhibit lipid-like densities.