Iodo--Conotoxin MI Selectively Binds the /
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- 作者：Siqin Luo and J. Michael McIntosh
- 刊名：Biochemistry
- 出版年：2004
- 出版时间：June 1, 2004
- 年：2004
- 卷：43
- 期：21
- 页码：6656 - 6662
- 全文大小：131K
- 年卷期：v.43,no.21(June 1, 2004)
- ISSN：1520-4995
文摘
The embryonic mouse muscle nicotinic acetylcholine receptor (nAChR) is a ligand-gatedion channel formed by 
1, 
1, 
, and 
subunits. The receptor contains two ligand binding sites at /and / subunit interfaces. [3H]Curare preferentially binds the / interface. We describe the synthesisand properties of a high-affinity iodinated ligand that selectively binds the / interface. An analogue of-conotoxin MI was synthesized with an iodine attached to Tyr-12 (iodo--MI). The analogue potentlyblocks the fetal mouse muscle subtype of nAChR expressed in Xenopus oocytes. It failed, however,to block 34, 42, or 7 nAChRs. Iodo--MI potently blocks the 11 but not the 11subunit combination expressed in Xenopus oocytes indicating selectivity for the / subunit interface.-Conotoxin MI was subsequently radioiodinated, and its properties were further evaluated. Saturationexperiments indicate that radioiodinated -conotoxin MI binds to TE671 cell homogenates with a Hillslope of 0.95 ± 0.0094. Kinetic studies indicate that the binding of [125I]-conotoxin MI is reversible(koff = 0.084 ± 0.0045 min-1); kon is 8.5 × 107 min-1 M-1. The calculated kd is 0.98 nM. This potencyis ~20-fold higher than the unmodified -MI peptide. Unlike [125I]-bungarotoxin, [125I]-conotoxin MIbinding to TE671 cell homogenates is fully displaceable by the small molecule antagonist d-tubocurarine.
1, 1, , and subunits. The receptor contains two ligand binding sites at /and / subunit interfaces. [3H]Curare preferentially binds the / interface. We describe the synthesisand properties of a high-affinity iodinated ligand that selectively binds the / interface. An analogue of-conotoxin MI was synthesized with an iodine attached to Tyr-12 (iodo--MI). The analogue potentlyblocks the fetal mouse muscle subtype of nAChR expressed in Xenopus oocytes. It failed, however,to block 34, 42, or 7 nAChRs. Iodo--MI potently blocks the 11 but not the 11subunit combination expressed in Xenopus oocytes indicating selectivity for the / subunit interface.-Conotoxin MI was subsequently radioiodinated, and its properties were further evaluated. Saturationexperiments indicate that radioiodinated -conotoxin MI binds to TE671 cell homogenates with a Hillslope of 0.95 ± 0.0094. Kinetic studies indicate that the binding of [125I]-conotoxin MI is reversible(koff = 0.084 ± 0.0045 min-1); kon is 8.5 × 107 min-1 M-1. The calculated kd is 0.98 nM. This potencyis ~20-fold higher than the unmodified -MI peptide. Unlike [125I]-bungarotoxin, [125I]-conotoxin MIbinding to TE671 cell homogenates is fully displaceable by the small molecule antagonist d-tubocurarine.