Structure Of TRPV1 Research Articles

Nanomaterial‐Mediated Modulation of TRPV1 Ion Channels for Biomedical Applications

AbstractTransient receptor potential vanilloid subtype 1 (TRPV1) is a nonselective cation channel involved in various physiological processes such as pain perception, thermoregulation, and inflammatory responses. Nanomaterials have emerged as precise tools to modulate TRPV1 activity, offering high spatiotemporal resolution and specificity. These nanomaterials act as transducers, responding to internal or external stimuli such as pH, light, electric, and magnetic fields to deliver modulatory agents like agonists, antagonists, heat, reactive species, and mechanical forces to TRPV1 channels. This strategy enables non‐invasive and targeted therapeutic interventions for diseases associated with TRPV1 dysfunction. In this review, recent advances are highlighted in nanomaterial‐mediated TRPV1 modulation and its biomedical applications. The TRPV1 structure and activation mechanisms, the integration of nanomaterials for effective TRPV1 modulation, and the required material properties are covered. Moreover, biomedical applications are discussed, including neurostimulation, neurological disorder therapies, cancer therapies, metabolic disease treatments, and cardiovascular disease interventions. Future research directions and challenges in this field are also proposed.

Thermoring basis for heat unfolding‐induced inactivation in TRPV1

AbstractTransient receptor potential vanilloid‐1 (TRPV1) is a capsaicin receptor that employs use‐dependent desensitization to protect highly evolved mammals from noxious heat damage in response to repeated or constant heat stimuli. However, the underlying structural factor or motif has not been precisely resolved. In this computational study, the graph theory‐based grid thermodynamic model was used to reveal how temperature‐dependent noncovalent interactions, as identified in the 3D structures of rat TRPV1, could develop a well‐organized fluidic grid‐like mesh network. This network features various topological grids constrained as thermosensitive rings that range in size from the biggest to the smallest, governing distinct structural and functional traits of the channel in response to different temperature degrees. After discovering that heat unfolding of three specific biggest grids, one in the closed state and two in the open state, respectively, causes the reversible activation at 43°C and thermal inactivation between 56°C and 61°C, a smaller random grid was also found to be responsible for irreversible inactivation and use‐dependent desensitization from the pre‐open closed state within the temperature range of 43°C–61°C. Thus, these two distinct inactivation pathways of TRPV1 may be involved in protecting those mammals against noxious heat damage.Key Points A perturbation at the protein–water interface was accompanied by partial heat or cold unfolding of the membrane protein. A reversible or irreversible gating transition of an ion channel may result from a specific or random interaction between two active sites, respectively. Kinetically driven protein aggregation was not the cause of thermodynamically trapped irreversible inactivation, but rather a later stage of partial heat‐induced unfolding.

Design and experimental validation of a new radiolabeled analog of N-(3-hydroxy-4-methoxy-phenyl-methyl) ferrocene-carboxamide (VFC) targeting the TRPV1 receptor

The TRPV1 receptor has been recognized to play a role in cancer development, being overexpressed in prostate, breast, lung, and colon cancers. Since TRPV1 activation promotes cancer cell proliferation, invasion, and migration, TRPV1 antagonists may show potential as anti-cancer agents. Capsaicin and capsaicinoids are phytochemicals derived from homovanillic acid found in abundance in chili peppers and responsible for its pungent properties. Capsaicin acts as a potent agonist of TRPV1 with recognized antineoplastic properties. Here, we employ computational approaches including molecular modeling and docking to refine the 3D structure of human TRPV1 and assess its interaction with the newly synthesized N-(3-hydroxy-4-methoxyphenylmethyl)ferrocenecarboxamide (VFC) and its cyclopentadienyl tricarbonyl rhenium and technetium analogs. Radiolabeling of VFC with 99mTc was achieved by double ligand exchange to afford 99mTc-VFC in high radiochemical purity and yield. Biodistribution studies in mice demonstrated preferential accumulation of 99mTc-VFC in the bladder, liver, kidney, and lung. These findings may contribute to developing efficient TRPV1-targeted radiotracers for molecular imaging of tumors by SPECT.

MOLECULAR INSIGHTS INTO PROPOXAZEPAM INTERACTION WITH TRPV1 RECEPTORS: A DOCKING ANALYSIS

Problem. Pain therapy has remained conceptually stagnant since the opioid crisis, which highlighted the dangers of treating pain with opioids. TRPV1 receptors and their ligands have emerged as promising targets for the management of various pain conditions, including inflammatory and neuropathic pain. Further investigation into the activation and desensitization mechanisms of TRPV1 receptors, as well as the exploration of novel ligands with enhanced selectivity and efficacy, holds the potential to alleviate the burden of chronic pain. Aim. The study of interactions of ligands with the TRPV1 receptor by molecular docking and analysis of components of these interactions. Methods. In order to study the binding energy of the TRPV1 receptor with the researched compounds 8GFA – Cryo-EM structure of human TRPV1 in complex with the analgesic drug SB-366791 was utilized. The docking and QSAR prediction were carried out with propoxazepam, its possible metabolite 3-hydroxopropoxazepam, diazepam, oxazepam, SB-366791, RTX, capsazepin, capsaicin. The docking and QSAR build model using the Maestro Schrödinger Suite. The main results. Propoxazepam has the necessary pharmacophoric features to bind with TRPV1 recepor: region A-benzene ring; region B – the amide group (NH-C=O); region C – alkoxy group. The docking score of propoxazepam (-7.30 kcal/mol) indicates a stronger interaction with the TRPV1 receptor compared to oxazepam (-6.82 kcal/mol), 3-hydroxopropoxazepam (-6.49 kcal/mol), and capsazepin (-6.39 kcal/mol). When considering the increase in the free energy of interactions, the ligands can be ranked as follows: SB-366791 > Capsaicin > RTX > Capsazepin > Propoxazepam > Diazepam > 3-hydroxopropoxazepam > Oxazepam. Propoxazepam establishes two hydrogen bonds: one involving the NH group of the amide (resulting in a hydrogen bond interaction with the linker-neck) and THR550 of the protein, and another between oxygen of the alkoxy group (hydrophobic tail) and TYR511 of the TRPV1 receptor. Ligands with confirmed effects on the TRPV1 receptor also engage in interactions with the protein by forming hydrogen bonds with the same amino acids as the benzodiazepines, namely THR550 and TYR511. Propoxazepam is predicted to have relatively low potency or affinity for TRPV1 (predicted pIC50 value of –1.115), but this value is higher than other ligands. Conclusions. As propoxazepam has the necessary pharmacophoric features of the pharmacophore model of TRPV1 ligands, it could theoretically bind with this receptor. Propoxazepam creates hydrogen bond with TYR511 of the TRPV1 receptor as referent ligand SB-366791. Propoxazepam exhibits one of the largest contributions of hydrogen bonds in the energy of interaction with the receptor. According to QSAR modelling, all studied compounds (3-hydroxopropoxazepam, diazepam, oxazepam, propoxazepam) have low pIC50 values, which could indicate a relatively low potency or affinity for TRPV1. But propoxazepam has the highest predicted pIC50, it means that the model predicts it to be the most potent among the compounds.

Opening of capsaicin receptor TRPV1 is stabilized equally by its four subunits

Capsaicin receptor TRPV1 is a nociceptor for vanilloid molecules, such as capsaicin and resiniferatoxin (RTX). Even though cryo-EM structures of TRPV1 in complex with these molecules are available, how their binding energetically favors the open conformation is not known. Here, we report an approach to control the number of bound RTX molecules (0-4) in functional rat TRPV1. The approach allowed direct measurements of each of the intermediate open states under equilibrium conditions at both macroscopic and single-molecule levels. We found that RTX binding to each of the four subunits contributes virtually the same activation energy, which we estimated to be 1.70 to 1.86kcal/mol and found to arise predominately from destabilizing the closed conformation. We further showed that sequential bindings of RTX increase open probability without altering single-channel conductance, confirming that there is likely a single open-pore conformation for TRPV1 activated by RTX.

Human TRPV1 structure and inhibition by the analgesic SB-366791

Pain therapy has remained conceptually stagnant since the opioid crisis, which highlighted the dangers of treating pain with opioids. An alternative addiction-free strategy to conventional painkiller-based treatment is targeting receptors at the origin of the pain pathway, such as transient receptor potential (TRP) ion channels. Thus, a founding member of the vanilloid subfamily of TRP channels, TRPV1, represents one of the most sought-after pain therapy targets. The need for selective TRPV1 inhibitors extends beyond pain treatment, to other diseases associated with this channel, including psychiatric disorders. Here we report the cryo-electron microscopy structures of human TRPV1 in the apo state and in complex with the TRPV1-specific nanomolar-affinity analgesic antagonist SB-366791. SB-366791 binds to the vanilloid site and acts as an allosteric hTRPV1 inhibitor. SB-366791 binding site is supported by mutagenesis combined with electrophysiological recordings and can be further explored to design new drugs targeting TRPV1 in disease conditions.

The past and future of transient receptor potential: A scientometric analysis.

Background:Thermoreceptors include TRPV1 and TRPM8. TRPV1 and TRPM8 are TRP channels. TRP ion channels are widely expressed in many different tissues and cell types, and are involved in different physiological processes. Research on the structure and basic physiological functions of TRPV1 is relatively perfect, and the relationship between the pathogenesis of other members of the TRP family and specific diseases and TRPV1 remains to be explored in depth.Methods:Articles regarding TRP were culled from the Web of Science Core Collection, and knowledge maps were generated using the CiteSpace software.Results:In total, 19,862 articles were included. The number of published articles on this topic has rapidly increased since 2000, with more than 1000 articles published per year by 2020. MAKOTO TOMINAGA was the author with the most articles. The countries with the most articles were the United States and China. However, the number of articles in the U.S. was 3 times that in China. The organizations that publish the most articles are Harvard University in the US and Seoul Natl University in South Korea. TRP and the pathogenesis of diseases, such as neuropathy and stroke, are hotspots of current research.Conclusion:To our knowledge, this is the first study to provide an overview of the literature on TRP. Research on TRPs is developing rapidly.

Genomic variance analysis to constrain mutagenesis studies of TRPV1 subunit interfaces

Protein structures help to identify key amino acids for further functional studies. However, mutagenesis of all interesting sites is time consuming, and a tool to inform our choices would be practical. After color-coding the residues of the rat TRPV1 structure (rTRPV1, 3j5p) according to the number of single nucleotide polymorphisms (SNPs) observed in gnomAD 2.1.1, we noted lower variability in buried amino acids of the ankyrin repeat domain (ARD). We hypothesized that amino acids critical to interactions across an interface between subunits would exhibit a similar reduced variability.

TRPV1: Role in Skin and Skin Diseases and Potential Target for Improving Wound Healing.

Skin is innervated by a multitude of sensory nerves that are important to the function of this barrier tissue in homeostasis and injury. The role of innervation and neuromediators has been previously reviewed so here we focus on the role of the transient receptor potential cation channel, subfamily V member 1 (TRPV1) in wound healing, with the intent of targeting it in treatment of non-healing wounds. TRPV1 structure and function as well as the outcomes of TRPV1-targeted therapies utilized in several diseases and tissues are summarized. In skin, keratinocytes, sebocytes, nociceptors, and several immune cells express TRPV1, making it an attractive focus area for treating wounds. Many intrinsic and extrinsic factors confound the function and targeting of TRPV1 and may lead to adverse or off-target effects. Therefore, a better understanding of what is known about the role of TRPV1 in skin and wound healing will inform future therapies to treat impaired and chronic wounds to improve healing.

Predicting lipid and ligand binding sites in TRPV1 channel by molecular dynamics simulation and machine learning.

As a key cellular sensor, the TRPV1 channel undergoes a gating transition from a closed state to an open state in response to many physical and chemical stimuli. This transition is regulated by small-molecule ligands including lipids and various agonists/antagonists, but the underlying molecular mechanisms remain obscure. Thanks to recent revolution in cryo-electron microscopy, a growing list of new structures of TRPV1 and other TRPV channels have been solved in complex with various ligands including lipids. Toward elucidating how ligand binding correlates with TRPV1 gating, we have performed extensive molecular dynamics simulations (with cumulative time of 20 μs), starting from high-resolution structures of TRPV1 in both the closed and open states. By comparing between the open and closed state ensembles, we have identified state-dependent binding sites for small-molecule ligands in general and lipids in particular. We further use machine learning to predict top ligand-binding sites as important features to classify the closed vs open states. The predicted binding sites are thoroughly validated by matching homologous sites in all structures of TRPV channels bound to lipids and other ligands, and with previous functional/mutational studies of ligand binding in TRPV1. Taken together, this study has integrated rich structural, dynamic, and functional data to inform future design of small-molecular drugs targeting TRPV1.

Molecular Docking Analysis of Ginger Active Compound on Transient Receptor Potential Cation Channel Subfamily V Member 1 (TRPV1)

Ginger had been reported to ameliorate painful diabetic neuropathy (PDN) in an animal model. Gingerol and shogaol were active compounds of ginger that potentially act on transient receptor potential cation channel subfamily V member 1 (TRPV1), a key receptor in PDN. This study aims to predict the binding of gingerol and shogaol to TRPV1 using an in silico model. The ligands of the docking study were 3 chemical compounds of each gingerol and shogaol, i.e. 6-shogaol, 8-shogaol, 10-shogaol, 6-gingerol, 8 gingerol and 10-gingerol. Capsaicin, a TRPV1 agonist, was used as a native ligand. The TRPV1 structure was taken from Protein Data Bank (ID 3J9J). The docking analysis was performed using Autodock Vina. The result showed that among the ginger active compounds, 6-shogaol had the strongest binding energy (-7.10 kcal/mol) to TRPV1. The 6-shogaol lacked the potential hydrogen bond to Ile265 of TRPV1 protein, which capsacin had. However, it's binding energy towards TRPV1 was not significantly different compared to capsaicin. Therefore, 6-shogaol had potential to be developed as a treatment for PDN.

Thermal Diffusion Pathways of TRPV1

TRPV1 is a polymodal receptor gated by vanilloid ligands, heat, voltage, lipids, protons, and peptides from venomous animals (Diaz-Franulic et al., 2017). The structure of TRPV1 in the closed and open configurations (Gao et al., 2016) has been resolved and may provide a frame work to identify the unknown structure(s) responsible of heat sensing in TRPV1 channels. Here, taking advantage of the known TRPV1 structures and using molecular simulations and the anisotropic thermal diffusion method (ATD), we evaluated the heat transfer pathways which could be responsible to guide the thermal energy from the environment to the channels gates. ATD is a non-equilibrium molecular dynamics simulation method that consists in introducing vibrational energy in each amino acid of the channel-forming protein and then to measure its impact in the kinetic energy of the remaining residues (Ota, N. and D.A. Agard, 2005). Thus, the ATD technique allows us to map the heat diffusion pathways in the TRPV1 channel and identify the amino acid involved in the allosteric communication between the TRPV1 channel structures absorbing energy from the surroundings and the channel gates. Here, we present a detailed analysis of the amino acids involved in well-defined thermal diffusion pathways, and we show that most of the thermal pathways identified converge into the hydrophobic gating ring formed by Ile679.

The Role of the Selectivity Filter in TRPV1 Channel Gating

The TRPV1 channel is a homotetrameric non-selective cation channel that functions in nociceptors as an integrator of external noxious stimuli and endogenous pro-inflammatory molecules. Although the binding sites for some TRPV1 modulators have been characterized, we still don’t understand how any of these stimuli influence ion conduction in this receptor. The identification and characterization of the regions that function as activation gates are therefore central to understanding the mechanisms of gating in the TRPV1 and other related channels. The structures of TRPV1 in open and closed states suggested that in addition to the intracellular gate formed by the pore-lining S6 helices, the selectivity filter could also function as an activation gate. The available structures of other TRP channels also support the existence of two gates, suggesting that it could be a conserved and unique feature of the TRP family. Here we set out to determine whether the selectivity filter of TRPV1 functions as a gate. We have substituted cysteines along the pore using a cysteineless channel background and assessed their accessibility to externally applied silver ions in both the open and the closed states in patch clamp recordings. First we found that external silver ions act as high-affinity, voltage-dependent, permeant blockers of the cysteineless TRPV1 channel, with a higher affinity for the closed state relative to the open state, suggesting that the filter does not function as a gate. We have provided further support for this hypothesis by showing that accessibility to external silver ions to a cysteine more intracellular than the filter is not affected by channel gating. It is possible that the filter adopts a non-conducting conformation under specific conditions yet to be identified, or that it constitutes a highly dynamic region of the receptor.

Decrypting the Heat Activation Mechanism of TRPV1 Channel by Molecular Dynamics Simulation

As a prototype cellular sensor, the TRPV1 cation channel undergoes a closed-to-open gating transition in response to various physical and chemical stimuli including noxious heat. Despite recent progress, the molecular mechanism of heat activation of TRPV1 gating remains enigmatic. Toward decrypting the structural basis of TRPV1 heat activation, we performed extensive molecular dynamics (MD) simulations (with cumulative simulation time of ∼11 μs) for the wildtype (WT) channel and a constitutively active double mutant at different temperatures (30°C, 60°C, and 72°C), starting from a high-resolution closed-channel structure of TRPV1 solved by cryo-electron microscopy (cryo-EM). In the WT simulations, we observed heat-activated conformational changes (e.g., expansion or contraction) in various key domains of TRPV1 (e.g., the S2-S3 and S4-S5 linkers) to prime the channel for gating. These conformational changes involve a number of dynamic hydrogen-bond interactions which were validated with previous mutational studies. Next, our mutant simulations observed channel opening following a series of conformational changes that propagate from the channel periphery to the channel pore via key intermediate domains (including the S2-S3 and S4-S5 linkers). The gating transition is accompanied by a large increase in the protein-water electrostatic interaction energy which supports the contribution of de-solvation of polar/charged residues to the temperature-sensitive TRPV1 gating. Taken together, our MD simulations and analyses offered new structural, dynamic, and energetic information to guide future mutagenesis and functional studies of TRPV1 channel and development of TRPV1-targeting drugs.

Decrypting the Heat Activation Mechanism of TRPV1 Channel by Molecular Dynamics Simulation

As a prototype cellular sensor, the TRPV1 cation channel undergoes a closed-to-open gating transition in response to various physical and chemical stimuli including noxious heat. Despite recent progress, the molecular mechanism of heat activation of TRPV1 gating remains enigmatic. Toward decrypting the structural basis of TRPV1 heat activation, we performed extensive molecular dynamics simulations (with cumulative simulation time of ∼11 μs) for the wild-type channel and a constitutively active double mutant at different temperatures (30, 60, and 72°C), starting from a high-resolution closed-channel structure of TRPV1 solved by cryo-electron microscopy. In the wild-type simulations, we observed heat-activated conformational changes (e.g., expansion or contraction) in various key domains of TRPV1 (e.g., the S2-S3 and S4-S5 linkers) to prime the channel for gating. These conformational changes involve a number of dynamic hydrogen-bond interactions that were validated with previous mutational studies. Next, our mutant simulations observed channel opening after a series of conformational changes that propagate from the channel periphery to the channel pore via key intermediate domains (including the S2-S3 and S4-S5 linkers). The gating transition is accompanied by a large increase in the protein-water electrostatic interaction energy, which supports the contribution of desolvation of polar/charged residues to the temperature-sensitive TRPV1 gating. Taken together, our molecular dynamics simulations and analyses offered, to our knowledge, new structural, dynamic, and energetic information to guide future mutagenesis and functional studies of the TRPV1 channels and development of TRPV1-targeting drugs.

Visualizing small molecule binding in membrane protein by single particle cryo-EM

In the last few years, major technological breakthroughs, particularly the development of new direct electron detection cameras and associated technologies, have enabled single particle cryogenic electron microscopy (cryo-EM) to become the technique of choice for high-resolution structure determination of many challenging biological macromolecules. Atomic structures of many membrane proteins, particularly ion channels, that are refractory to crystallization have now determined by this method, including our own work of determining the atomic structures of TRPV1, TRPA1 and TPRM4. A recent technological development in single particle cryo-EM of integral membrane protein is to enable atomic structure determination of integral membrane proteins in a native or native-like lipid bilayer environment. In many recent cryo-EM studies, integral membrane proteins are reconstituted into native like lipid bilayer environment, using various lipid-protein nanoparticle techniques including lipid nanodisc, saposin based Salipro nanoparticles, or the styrene-maleic acid (SMA) copolymer based native lipid nanodisc. By reconstituting integral membrane proteins into artificial or near native lipid bi-layer environment, it is now feasible to use single particle cryo-EM for studying specific protein - lipid interactions, and to visualize binding of small pharmacological substances in membrane protein in lipid environment.

Heat activation is intrinsic to the pore domain of TRPV1

The TRPV1 channel is a sensitive detector of pain-producing stimuli, including noxious heat, acid, inflammatory mediators, and vanilloid compounds. Although binding sites for some activators have been identified, the location of the temperature sensor remains elusive. Using available structures of TRPV1 and voltage-activated potassium channels, we engineered chimeras wherein transmembrane regions of TRPV1 were transplanted into the Shaker Kv channel. Here we show that transplanting the pore domain of TRPV1 into Shaker gives rise to functional channels that can be activated by a TRPV1-selective tarantula toxin that binds to the outer pore of the channel. This pore-domain chimera is permeable to Na+, K+, and Ca2+ ions, and remarkably, is also robustly activated by noxious heat. Our results demonstrate that the pore of TRPV1 is a transportable domain that contains the structural elements sufficient for activation by noxious heat.

Model-based local density sharpening of cryo-EM maps.

Atomic models based on high-resolution density maps are the ultimate result of the cryo-EM structure determination process. Here, we introduce a general procedure for local sharpening of cryo-EM density maps based on prior knowledge of an atomic reference structure. The procedure optimizes contrast of cryo-EM densities by amplitude scaling against the radially averaged local falloff estimated from a windowed reference model. By testing the procedure using six cryo-EM structures of TRPV1, β-galactosidase, γ-secretase, ribosome-EF-Tu complex, 20S proteasome and RNA polymerase III, we illustrate how local sharpening can increase interpretability of density maps in particular in cases of resolution variation and facilitates model building and atomic model refinement.

Side chain flexibility and coupling between the S4-S5 linker and the TRP domain in thermo-sensitive TRP channels: Insights from protein modeling.

The transient receptor potential (TRP) superfamily is subdivided into several subfamilies on the basis of sequence similarity, which is highly heterogeneous but shows a molecular architecture that resembles the one present in members of the Kv channel superfamily. Because of this diversity, they produce a large variety of channels with different gating and permeability properties. Elucidation of these particular features necessarily requires comparative studies based on structural and functional data. The present study aims to compilate, analyze, and determine, in a coherent way, the relationship between intrinsic side-chain flexibility and the allosteric coupling in members of the TRPV, TRPM, and TRPC families. Based on the recently determined structures of TRPV1 and TRPV2, we have generated protein models for single subunits of TRPV5, TRPM8, and TRPC5 channels. With these models, we focused our attention on the apparently crucial role of the GP dipeptide at the center of the S4-S5 linker and discussed its role in the interaction with the TRP domain, specifically with the highly-conserved Trp during this coupling. Our analysis suggests an important role of the S4-S5L flexibility in the thermosensitivity, where heat-activated channels possess rigid S4-S5 linkers, whereas cold-activated channels have flexible ones. Finally, we also present evidence of the key interaction between the conserved Trp residue of the TRP box and of several residues in the S4-S5L, importantly the central Pro. Proteins 2017; 85:630-646. © 2016 Wiley Periodicals, Inc.

Toward Elucidating the Heat Activation Mechanism of the TRPV1 Channel Gating by Molecular Dynamics Simulation

As a key cellular sensor, the TRPV1 cation channel undergoes a gating transition from a closed state to an open state in response to various physical and chemical stimuli including noxious heat. Despite years of study, the heat activation mechanism of TRPV1 gating remains enigmatic at the molecular level. Toward elucidating the structural and energetic basis of TRPV1 gating, we have performed extensive molecular dynamics (MD) simulations (with cumulative simulation time of 3 microseconds), starting from the high-resolution closed and open structures of TRPV1 solved by cryo-electron microscopy. In the closed-state simulations at 30°C, we observed a stably closed channel constricted at the lower gate (near residue I679), while the upper gate (near residues G643 and M644) is dynamic and undergoes flickery opening/closing. In the open-state simulations at 60°C, we found higher conformational variation consistent with a large entropy increase required for the heat activation, and both the lower and upper gates are dynamic with transient opening/closing. Through ensemble-based structural analyses of the closed state vs. the open state, we revealed pronounced closed-to-open conformational changes involving the membrane proximal domain (MPD) linker, the outer pore, and the TRP helix, which are accompanied by breaking/forming of a network of closed/open-state specific hydrogen bonds. By comparing the closed-state simulations at 30°C and 60°C, we observed heat-activated conformational changes in the MPD linker, the outer pore, and the TRP helix that resemble the closed-to-open conformational changes, along with partial formation of the open-state specific hydrogen bonds. Some of the residues involved in the above key hydrogen bonds were validated in comparison with previous mutational studies. Taken together, our MD simulations have offered rich structural and dynamic details beyond the static structures of TRPV1, and promising targets for future mutagenesis and functional studies of the TRPV1 channel.