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J Biol Chem. 2019 Dec 20;294(51):19589-19603. doi: 10.1074/jbc.RA119.009959. Epub 2019 Nov 14.

Altered allostery of the left flipper domain underlies the weak ATP response of rat P2X5 receptors.

Author information

1
Institute of Medical Sciences and Department of Pharmacology and Physiology, Shanghai Jiao Tong University School of Medicine, Shanghai 200025, China.
2
Putuo District Center Hospital, Shanghai University of Chinese Traditional Medicine, Shanghai 200026, China.
3
Department of Integrative Biology and Pharmacology, McGovern Medical School, University of Texas Health Science Center, Houston, Texas 77030.
4
State Key Laboratory for Chemistry and Molecular Engineering of Medicinal Resources, School of Chemistry and Pharmacy, Guangxi Normal University, Guilin 541004, China.
5
College of Bioscience and Biotechnology, Hunan Agricultural University, Changsha 410128, China.
6
State Key Laboratory for Chemistry and Molecular Engineering of Medicinal Resources, School of Chemistry and Pharmacy, Guangxi Normal University, Guilin 541004, China hshwang2002@163.com.
7
Department of Basic Medicine and Clinical Pharmacy, China Pharmaceutical University, Nanjing 210009, China crguocpu@126.com.
8
Institute of Medical Sciences and Department of Pharmacology and Physiology, Shanghai Jiao Tong University School of Medicine, Shanghai 200025, China yuye@shsmu.edu.cn.
9
Department of Basic Medicine and Clinical Pharmacy, China Pharmaceutical University, Nanjing 210009, China.

Abstract

Although the extracellular ATP-gated cation channel purinergic receptor P2X5 is widely expressed in heart, skeletal muscle, and immune and nervous systems in mammals, little is known about its functions and channel-gating activities. This lack of knowledge is due to P2X5's weak ATP responses in several mammalian species, such as humans, rats, and mice. WT human P2X5 (hP2X5Δ328-349) does not respond to ATP, whereas a full-length variant, hP2X5 (hP2X5-FL), containing exon 10 encoding the second hP2X5 transmembrane domain (TM2), does. However, although rat P2X5 (rP2X5) has a full-length TM2, ATP induces only weak currents in rP2X5, which prompted us to investigate the mechanism underlying this small ATP response. Here, we show that single replacements of specific rP2X5 residues with the corresponding residues in hP2X5 (S191F or F195H) significantly enhance the current amplitude of rP2X5. Using a combination of engineered disulfide cross-linking, single-channel recording, and molecular modeling, we interrogated the effects of S191F and F195H substitutions on the allostery of the left flipper (LF) domain. On the basis of our findings, we propose that the bound ATP-induced distinct allostery of the LF domain with that of other functional subtypes has caused the weak ATP response of rP2X5 receptors. The findings of our study provide the prerequisite for future transgenic studies on the physiological and pathological functions of P2X5 receptors.

KEYWORDS:

ATP-evoked current; ATP-gated ion channel; LF domain; P2X5 receptors; allosteric regulation; conformational change; gating; left flipper domain; molecular dynamics; purinergic receptor

PMID:
31727741
PMCID:
PMC6926468
[Available on 2020-12-20]
DOI:
10.1074/jbc.RA119.009959

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