Structure and Function of the ThermoTRP Channel Pore

Zheng, Jie; Ma, Linlin

doi:10.1016/b978-0-12-800181-3.00009-9

Cited by 30 publications

(36 citation statements)

References 87 publications

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“…We further observed that the head of resiniferatoxin forms hydrogen bonds with E571 on S4-S5 linker and S513 on S3 (Fig. 6A, dashed lines), which glues the S4-S5 linker to the anticipated stationary S1-S4 domain (30). Hence, although different parts of vanilloids are involved in interactions with TRPV1, the underlying activation mechanism appears to be analogous for capsaicin and resiniferatoxin (Fig.…”

Section: Trpv1mentioning

confidence: 91%

“…Conformational change in S4 is coupled to the movement of the S4-S5 linker to induce a conformational change in S6 interacting with the linker, leading to the opening of the activation gate (35)(36)(37). Although the S1-S4 domains of TRP channels are structurally similar to that of Kv channels (11,18,19,38,39), no positively charged residue exists in the S4 of TRPV channels so this domain is expected to remain stationary upon membrane depolarization or the presence of other activators (30). This allows capsaicin or resiniferatoxin to first secure itself to the stationary S1-S4 domain, and then stabilize the outward conformation of the S4-S5 linker to mediate S6 movements leading to channel activation (6,40).…”

Section: Discussionmentioning

confidence: 99%

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Rational design and validation of a vanilloid-sensitive TRPV2 ion channel

Yang

Yarov‐Yarovoy

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

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Vanilloids activation of TRPV1 represents an excellent model system of ligand-gated ion channels. Recent studies using cryo-electron microcopy (cryo-EM), computational analysis, and functional quantification revealed the location of capsaicin-binding site and critical residues mediating ligand-binding and channel activation. Based on these new findings, here we have successfully introduced high-affinity binding of capsaicin and resiniferatoxin to the vanilloid-insensitive TRPV2 channel, using a rationally designed minimal set of four point mutations (F467S-S498F-L505T-Q525E, termed TRPV2_Quad). We found that binding of resiniferatoxin activates TRPV2_Quad but the ligand-induced open state is relatively unstable, whereas binding of capsaicin to TRPV2_Quad antagonizes resiniferatoxin-induced activation likely through competition for the same binding sites. Using Rosettabased molecular docking, we observed a common structural mechanism underlying vanilloids activation of TRPV1 and TRPV2_Quad, where the ligand serves as molecular "glue" that bridges the S4-S5 linker to the S1-S4 domain to open these channels. Our analysis revealed that capsaicin failed to activate TRPV2_Quad likely due to structural constraints preventing such bridge formation. These results not only validate our current working model for capsaicin activation of TRPV1 but also should help guide the design of drug candidate compounds for this important pain sensor.on channels constitute the second largest family of drug targets for therapeutics (1-3); therefore, understanding their gating mechanisms by small-molecule ligands is critical for both basic and translational research. Capsaicin activation of the pain-sensing transient receptor potential vanilloid 1 (TRPV1) ion channel represents an outstanding model system for understanding liganddependent gating process (4), because capsaicin not only potently activates the channel with a submicromolar EC 50 (5) but also effectively stabilizes the channel at high open probability (6-8). Moreover, capsaicin activation is highly selective for TRPV1 channel (5). Previous mutagenesis (9, 10) and cryo-EM studies (11,12) have shown that capsaicin binds near the third (S3) and fourth (S4) transmembrane segments of TRPV1 (Fig. 1A). Based on the high-resolution cryo-EM structures, we have used a combination of computational and functional assays to reveal that capsaicin takes a "tail-up, head-down" configuration in the ligand-binding pocket (6) (Fig. 1B). The aliphatic tail forms extensive but nonspecific van der Waals (VDW) interactions with residues lining the binding pocket, whereas the vanillyl group (head) and the amide group (neck) of capsaicin form a hydrogen bond with E571 on S4-S5 linker and T551 on S4, respectively. To activate TRPV1, VDW interactions first secure capsaicin in the pocket and the neck of capsaicin forms a hydrogen bond with T551. This is followed by the formation of another hydrogen bond between the head and E571, which stabilizes the outward conformation of S4-S5 linker to open the chann...

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Section: Trpv1mentioning

confidence: 91%

Section: Discussionmentioning

confidence: 99%

Rational design and validation of a vanilloid-sensitive TRPV2 ion channel

Yang

Yarov‐Yarovoy

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

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“…Stabilization of TRPV1 for cryo-EM studies required deletion of the pore turret region 5,6 , while in the present work we employed full-length TRPV2 where the turret region is intact, although poorly resolved due to its flexibility. Homology modelling predicts that deletion of the turret region in the TRPV1 structures would alter the pore region architecture 34 , thus possibly accounting for some of the observed differences in pore architecture between TRPV1 and TRPV2 (Fig. 2).…”

Section: Resultsmentioning

confidence: 99%

“…Specifically, a B30 amino acid loop connecting S5 with the pore helix, known as the pore turret ( Fig. 4a), has been implicated in channel gating, large organic cation permeation and pore dilation 33,34 . Mutation of key residues within the TRPV1 pore turret (Gly 618 and Ser 629), which are conserved in TRPV2 (Gly 582 and Ser 592; Fig.…”

Section: Resultsmentioning

confidence: 99%

Structure of the Full-Length TRPV2 Channel by Cryo-EM

et al. 2016

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Transient receptor potential (TRP) proteins form a superfamily Ca 2 þ -permeable cation channels regulated by a range of chemical and physical stimuli. Structural analysis of a 'minimal' TRP vanilloid subtype 1 (TRPV1) elucidated a mechanism of channel activation by agonists through changes in its outer pore region. Though homologous to TRPV1, other TRPV channels (TRPV2-6) are insensitive to TRPV1 activators including heat and vanilloids. To further understand the structural basis of TRPV channel function, we determined the structure of full-length TRPV2 at B5 Å resolution by cryo-electron microscopy. Like TRPV1, TRPV2 contains two constrictions, one each in the pore-forming upper and lower gates. The agonist-free full-length TRPV2 has wider upper and lower gates compared with closed and agonist-activated TRPV1. We propose these newly revealed TRPV2 structural features contribute to diversity of TRPV channels.

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“…Noticeably, recent studies using centipede and spider toxins 6,21 , Mg 2+ 49,50 , and Na + 51 provided fresh evidence that, together with existing findings (summarized in ref. 52), suggests that the outer pore region is involved in the heat activation process. In the present study, we found that F430_3aa and E693_8aa, with almost completely decoupled Ca 2+ -dependent desensitization, exhibited nearly normal heat activation.…”

Section: Discussionmentioning

confidence: 99%

Exploring Functional Roles of TRPV1 Intracellular Domains with Unstructured Peptide-Insertion Screening

Yang

et al. 2017

Biophysical Journal

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TRPV1 is a polymodal nociceptor for diverse physical and chemical stimuli that interact with different parts of the channel protein. Recent cryo-EM studies revealed detailed channel structures, opening the door for mapping structural elements mediating activation by each stimulus. Towards this goal, here we have combined unstructured peptide-insertion screening (UPS) with electrophysiological and fluorescence recordings to explore structural and functional roles of the intracellular regions of TRPV1 in mediating various activation stimuli. We found that most of the tightly packed protein regions did not tolerate structural perturbation by UPS when tested, indicating that structural integrity of the intracellular region is critical. In agreement with previous reports, Ca 2+ -dependent desensitization is strongly dependent on both intracellular N-and C-terminal domains; insertions of an unstructured peptide between these domains and the transmembrane core domain nearly eliminated Ca 2+ -dependent desensitization. In contrast, channel activations by capsaicin, low pH, divalent cations, and even heat are mostly intact in mutant channels containing the same insertions. These observations suggest that the transmembrane core domain of TRPV1, but not the intracellular domains, is responsible for sensing these stimuli.TRPV1 ion channel can be directly activated or modulated by capsaicin 1 , heat 1 , extracellular proton 2 , divalent cations 3 , Na + 4 , peptide toxins from spider 5 , centipede 6 , and scorpion 7 , membrane depolarization 8 , intracellular Ca 2+ 9 , and many other factors 10 . A noticeable feature of TRPV1 polymodal activation emerging from biophysical investigations is the existence of non-overlapping activation pathways [11][12][13] . However, except for capsaicin 14-19 , proton 11,20 , and animal toxins 6,21 , the channel structures that sense activation stimuli are poorly defined. Recent cryo-EM studies revealed that the transmembrane core domain of TRPV1 resembles that of tetrameric cation channels, with six transmembrane helical segments surrounding a centrally located ion permeation pore 22,23 . The partially resolved intracellular N-and C-terminal domains contain special structures such as the ankyrin-like repeat domains and the TRP domain that likely play crucial structural and/or functional roles 10 . Arrangement of intracellular domains has been investigated using fluorescence resonance energy transfer 24 . A major task for TRPV1 study in the post-structure era is to assign functional roles to individual structure domains.The unstructured peptide-insertion screening (UPS) strategy had been used to rapidly and reliably obtain information on structure-function relationships in an ion channel even before detailed protein structures were available. In one effective application, isolating the pore domain of yeast TRPY1 channel from the intracellular Ca 2+ -sensing domain with unstructured peptides demonstrated that the pore domain was mechanosensitive 25 . Similarly, unstructured peptide insertion...

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Structure and Function of the ThermoTRP Channel Pore

Cited by 30 publications

References 87 publications

Rational design and validation of a vanilloid-sensitive TRPV2 ion channel

Rational design and validation of a vanilloid-sensitive TRPV2 ion channel

Structure of the Full-Length TRPV2 Channel by Cryo-EM

Exploring Functional Roles of TRPV1 Intracellular Domains with Unstructured Peptide-Insertion Screening

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