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Title: Distinct properties of Ca 2+-calmodulin binding to N- and C-terminal regulatory regions of the TRPV1 channel

Abstract

Transient receptor potential (TRP) vanilloid 1 (TRPV1) is a molecular pain receptor belonging to the TRP superfamily of nonselective cation channels. As a polymodal receptor, TRPV1 responds to heat and a wide range of chemical stimuli. The influx of calcium after channel activation serves as a negative feedback mechanism leading to TRPV1 desensitization. The cellular calcium sensor calmodulin (CaM) likely participates in the desensitization of TRPV1. Two CaM-binding sites are identified in TRPV1: the N-terminal ankyrin repeat domain (ARD) and a short distal C-terminal (CT) segment. Here, we present the crystal structure of calcium-bound CaM (Ca 2+–CaM) in complex with the TRPV1-CT segment, determined to 1.95-Å resolution. The two lobes of Ca 2+–CaM wrap around a helical TRPV1-CT segment in an antiparallel orientation, and two hydrophobic anchors, W787 and L796, contact the C-lobe and N-lobe of Ca 2+–CaM, respectively. This structure is similar to canonical Ca 2+–CaM-peptide complexes, although TRPV1 contains no classical CaM recognition sequence motif. Using structural and mutational studies, we established the TRPV1 C terminus as a high affinity Ca 2+–CaM-binding site in both the isolated TRPV1 C terminus and in full-length TRPV1. Although a ternary complex of CaM, TRPV1-ARD, and TRPV1-CT had previously been postulated, wemore » found no biochemical evidence of such a complex. In electrophysiology studies, mutation of the Ca 2+–CaM-binding site on TRPV1-ARD abolished desensitization in response to repeated application of capsaicin, whereas mutation of the Ca 2+–CaM-binding site in TRPV1-CT led to a more subtle phenotype of slowed and reduced TRPV1 desensitization. In summary, our results show that the TRPV1-ARD is an important mediator of TRPV1 desensitization, whereas TRPV1-CT has higher affinity for CaM and is likely involved in separate regulatory mechanisms.« less

Authors:
; ;  [1]
  1. Harvard
Publication Date:
Research Org.:
Argonne National Lab. (ANL), Argonne, IL (United States). Advanced Photon Source (APS)
Sponsoring Org.:
USDOE Office of Science (SC)
OSTI Identifier:
1054358
Resource Type:
Journal Article
Journal Name:
Journal of General Physiology
Additional Journal Information:
Journal Volume: 140; Journal Issue: (5) ; 2012; Journal ID: ISSN 0022-1295
Country of Publication:
United States
Language:
ENGLISH

Citation Formats

Lau, Sze-Yi, Procko, Erik, and Gaudet, Rachelle. Distinct properties of Ca2+-calmodulin binding to N- and C-terminal regulatory regions of the TRPV1 channel. United States: N. p., 2012. Web. doi:10.1085/jgp.201210810.
Lau, Sze-Yi, Procko, Erik, & Gaudet, Rachelle. Distinct properties of Ca2+-calmodulin binding to N- and C-terminal regulatory regions of the TRPV1 channel. United States. doi:10.1085/jgp.201210810.
Lau, Sze-Yi, Procko, Erik, and Gaudet, Rachelle. Thu . "Distinct properties of Ca2+-calmodulin binding to N- and C-terminal regulatory regions of the TRPV1 channel". United States. doi:10.1085/jgp.201210810.
@article{osti_1054358,
title = {Distinct properties of Ca2+-calmodulin binding to N- and C-terminal regulatory regions of the TRPV1 channel},
author = {Lau, Sze-Yi and Procko, Erik and Gaudet, Rachelle},
abstractNote = {Transient receptor potential (TRP) vanilloid 1 (TRPV1) is a molecular pain receptor belonging to the TRP superfamily of nonselective cation channels. As a polymodal receptor, TRPV1 responds to heat and a wide range of chemical stimuli. The influx of calcium after channel activation serves as a negative feedback mechanism leading to TRPV1 desensitization. The cellular calcium sensor calmodulin (CaM) likely participates in the desensitization of TRPV1. Two CaM-binding sites are identified in TRPV1: the N-terminal ankyrin repeat domain (ARD) and a short distal C-terminal (CT) segment. Here, we present the crystal structure of calcium-bound CaM (Ca2+–CaM) in complex with the TRPV1-CT segment, determined to 1.95-Å resolution. The two lobes of Ca2+–CaM wrap around a helical TRPV1-CT segment in an antiparallel orientation, and two hydrophobic anchors, W787 and L796, contact the C-lobe and N-lobe of Ca2+–CaM, respectively. This structure is similar to canonical Ca2+–CaM-peptide complexes, although TRPV1 contains no classical CaM recognition sequence motif. Using structural and mutational studies, we established the TRPV1 C terminus as a high affinity Ca2+–CaM-binding site in both the isolated TRPV1 C terminus and in full-length TRPV1. Although a ternary complex of CaM, TRPV1-ARD, and TRPV1-CT had previously been postulated, we found no biochemical evidence of such a complex. In electrophysiology studies, mutation of the Ca2+–CaM-binding site on TRPV1-ARD abolished desensitization in response to repeated application of capsaicin, whereas mutation of the Ca2+–CaM-binding site in TRPV1-CT led to a more subtle phenotype of slowed and reduced TRPV1 desensitization. In summary, our results show that the TRPV1-ARD is an important mediator of TRPV1 desensitization, whereas TRPV1-CT has higher affinity for CaM and is likely involved in separate regulatory mechanisms.},
doi = {10.1085/jgp.201210810},
journal = {Journal of General Physiology},
issn = {0022-1295},
number = (5) ; 2012,
volume = 140,
place = {United States},
year = {2012},
month = {11}
}