Tài liệu Báo cáo khoa học: a-Conotoxins as tools for the elucidation of structure and function of

MINIREVIEW

a-Conotoxins as tools for the elucidation of structure and function

of neuronal nicotinic acetylcholine receptor subtypes

Annette Nicke1

, Susan Wonnacott2 and Richard J. Lewis3

1

Max Planck-Institute for Brain Research, Frankfurt, Germany; 2

Department of Biology & Biochemistry, University of Bath, UK; 3

Institute for Molecular Bioscience, University of Queensland, Brisbane, Australia

Cone snails comprise
500 species of venomous molluscs,

which have evolved the ability to generate multiple toxins

with varied and often exquisite selectivity. One class,

the a-conotoxins, is proving to be a powerful tool for

the differentiation of nicotinic acetylcholine receptors

(nAChRs). These comprise a large family of complex

subtypes, whose significance in physiological functions and

pathological conditions is increasingly becoming apparent.

After a short introduction into the structure and diversity of

nAChRs, this overview summarizes the identification and

characterization of a-conotoxins with selectivity for neur￾onal nAChR subtypes and provides examples of their use in

defining the compositions and function of neuronal nAChR

subtypes in native vertebrate tissues.

Keywords: a-conotoxins; neuronal nicotinic acetylcholine

receptor subtypes; pharmacology; venom peptides; Xenopus

oocytes.

Neuronal nicotinic acetylcholine receptors

The nicotinic acetylcholine receptor family

The nicotinic acetylcholine receptor (nAChR) at the neuro￾muscular junction was first described as the
receptive

1substance in Langley’s historic experiments which lead to

the formulation of the receptor concept [1]. nAChRs have

been amongst the earliest receptors to be investigated

by pharmacological, biochemical, electrophysiological and

molecular biological approaches, and to date represent one

of the most intensively investigated membrane proteins.

While the identification and pharmacological distinction of

nAChR subtypes at the neuromuscular endplate (causing

muscle contraction) and those in sympathetic and para￾sympathetic ganglia (mediating neurotransmission) was

made relatively early, the existence of nAChRs in the brain

was controversial until cloning of the first neuronal nAChR

isoforms in the mid 1980s [2,3]. nAChRs are ligand-gated

ion channels that belong to the Cys-loop receptor super￾family which includes GABAA, glycine and 5HT3 neuro￾transmitter receptors.

The electric organs of the electric ray Torpedo and

eel Electrophorus provided a rich source of nAChRs that

facilitated their early structural characterization. The

nAChR from Torpedo californica is the best investigated

ligand-gated ion channel so far and considered as a

prototype. By electron microscopy techniques [4], high

resolution images down to 4 A˚ have been obtained from

semicrystalline arrays of this receptor in Torpedo mem￾branes. These studies revealed the pentameric quaternary

structure of this protein (Fig. 1) and have provided valuable

information about the channel architecture and dimensions.

A deeper insight into the molecular structure, in particular

the acetylcholine (ACh) binding pocket, has become

available after crystallization of an ACh binding protein,

which has high homology to the extracellular domain of

2the nAChR (Fig. 1) [5,6]. The Torpedo nAChR and the

nAChR in embryonic vertebrate muscle share the same

heteropentameric structure composed of four homologous

subunits which are arranged in the order a1ca1db1 around

the central ion-conducting channel [7,8] (Fig. 2A). In

addition, 11 nAChR subunits (a2–a7, a9, a10, b2–b4) have

been cloned from neuronal and sensory mammalian tissues.

A mammalian homologue of the avian a8 subunithas not

been found [2,3,9].

Subunit assembly of neuronal nAChRs

The a7, a8 and a9 subunits represent a subclass of neuronal

nAChRs that is able to form functional homomeric

channels upon heterologous expression [2,3]. Coexpression

of a7 and a8, as well as of a9 and the highly homologous

a10 subunit [10] has been shown to generate heteromeric

channels with properties distinct from those of the respective

homopentamers. The association of a7 wit h b subunits in

native nAChRs has been controversial [11]. The a2, a3, a4

and a6 subunits require coexpression of atleastone b (b2 or

b4) subunit to form functional channels [2,3,9]. However,

pairwise combinations of the a6 wit h t he b2 or b4 subunit

resulted in protein aggregation or very inefficient expression

of functional channels [12], indicating that at least two other

subunits are required for effective channel formation. In

Correspondence to A. Nicke, Max Planck-Institute for Brain Research,

Deutschordenstr. 46, D-60528 Frankfurt, Germany.

Fax: + 49 69 96769 441, Tel.: + 49 69 96769 262,

E-mail: [email protected]

Abbreviations: ACh, acetylcholine; nAChR, nicotinic acetylcholine

receptor; a-BTX, a-bungarotoxin; all a-conotoxins are abbreviated,

e.g. MII instead of a-conotoxin MII.

(Received 22 January 2004, revised 17 March 2004,

accepted 6 April 2004)

Eur. J. Biochem. 271, 2305–2319 (2004)
FEBS 2004 doi:10.1111/j.1432-1033.2004.04145.x