Preparation and characterization of a monoclonal antibody against the refolded and functional extracellular domain of rat P2X4 receptor

被引：11

作者：

Igawa, Tatsuhiro ^{[1
]}

Higashi, Sadayuki ^{[1
]}

Abe, Yoshito ^{[1
]}

Ohkuri, Takatoshi ^{[2
]}

Tanaka, Hiroyuki ^{[3
]}

Morimoto, Satoshi ^{[3
]}

Yamashita, Tomohiro ^{[4
]}

Tsuda, Makoto ^{[4
]}

Inoue, Kazuhide ^{[4
]}

Ueda, Tadashi ^{[1
]}

机构：

[1] Kyushu Univ, Dept Prot Struct Funct & Design, Grad Sch Pharmaceut Sci, Higashi Ku, Fukuoka 8128582, Japan

[2] Sojo Univ, Fac Pharmaceut Sci, Nishi Ku, Kumamoto 8600082, Japan

[3] Kyushu Univ, Dept Pharmacognosy, Grad Sch Pharmaceut Sci, Higashi Ku, Fukuoka 8128582, Japan

[4] Kyushu Univ, Dept Mol & Syst Pharmacol, Grad Sch Pharmaceut Sci, Higashi Ku, Fukuoka 8128582, Japan

来源：

JOURNAL OF BIOCHEMISTRY | 2013年 / 153卷 / 03期

关键词：

monoclonal antibody; neuropathic pain; P2X4 purinergic receptor; refolding; P2X(4) RECEPTORS; NEUROPATHIC PAIN; ION-CHANNEL; ATP; ACTIVATION; PROTEINS; FORCE; GLIA;

D O I：

10.1093/jb/mvs143

中图分类号：

Q5 [生物化学]; Q7 [分子生物学];

学科分类号：

071010 ; 081704 ;

摘要：

The P2X4 purinergic receptor is a key molecule in neuropathic pain, particularly in the allodynia after peripheral nerve injury. We therefore sought to establish an anti-P2X4 receptor monoclonal antibody that would be useful for detection and characterization of the P2X4 receptor. We first prepared the refolded extracellular domain of the rat P2X4 receptor expressed in Escherichia coli. Then, we established a B-cell hybridoma producing the monoclonal antibody for the head domain of the rat P2X4 receptor with strict recognition, including S-S bond formation. In addition, we succeeded in the detection and immune precipitation of rat P2X4 receptor molecules on cultured cells. As the antibody specifically binds to the rat P2X4 receptor molecule, it is expected that the established monoclonal antibody will be applicable as a tool for detecting increasing expression levels of the P2X4 receptor molecule accompanied with increasing intensity of neuropathic pain.

引用

页码：275 / 282

页数：8

共 21 条

[1] BDNF from microglia causes the shift in neuronal anion gradient underlying neuropathic pain
Coull, JAM
Beggs, S
Boudreau, D
Boivin, D
Tsuda, M
Inoue, K
Gravel, C
Salter, MW
De Koninck, Y
[J]. NATURE, 2005, 438 (7070) : 1017 - 1021
[2] Structural interpretation of P2X receptor mutagenesis studies on drug action
Evans, Richard J.
[J]. BRITISH JOURNAL OF PHARMACOLOGY, 2010, 161 (05) : 961 - 971
[3] Galfre G, 1981, Methods Enzymol, V73, P3
[4] ANTIBODY-PRODUCTION BY HYBRIDOMAS
GODING, JW
[J]. JOURNAL OF IMMUNOLOGICAL METHODS, 1980, 39 : 285 - 308
[5] Molecular mechanism of ATP binding and ion channel activation in P2X receptors
Hattori, Motoyuki
Gouaux, Eric
[J]. NATURE, 2012, 485 (7397) : 207 - U91
[6] P2X4 receptor is a glycosylated cardiac receptor mediating a positive inotropic response to ATP
Hu, B
Senkler, C
Yang, A
Soto, F
Liang, BT
[J]. JOURNAL OF BIOLOGICAL CHEMISTRY, 2002, 277 (18) : 15752 - 15757
[7] Crystal structure of the ATP-gated P2X4 ion channel in the closed state
Kawate, Toshimitsu
Michel, Jennifer Carlisle
Birdsong, William T.
Gouaux, Eric
[J]. NATURE, 2009, 460 (7255) : 592 - U55
[8] Crossing the pain barrier: P2 receptors as targets for novel analgesics
Kennedy, C
Assis, TS
Currie, AJ
Rowan, EG
[J]. JOURNAL OF PHYSIOLOGY-LONDON, 2003, 553 (03): : 683 - 694
[9] Khakh BS, 2001, PHARMACOL REV, V53, P107
[10] Clustal W and clustal X version 2.0
Larkin, M. A.
Blackshields, G.
Brown, N. P.
Chenna, R.
McGettigan, P. A.
McWilliam, H.
Valentin, F.
Wallace, I. M.
Wilm, A.
Lopez, R.
Thompson, J. D.
Gibson, T. J.
Higgins, D. G.
[J]. BIOINFORMATICS, 2007, 23 (21) : 2947 - 2948

← 1 2 3 →