Identification of an amino acid sequence within GABA(A) receptor beta(3) subunits that is important for receptor assembly

GABA(A) receptors are chloride ion channels that can be opened by GABA, the most important inhibitory transmitter in the CNS. In the mammalian brain the majority of these pentameric receptors is composed of two alpha, two beta and one gamma subunit. To achieve the correct order of subunits around the pore, each subunit must form specific contacts via its plus (+) and minus (-) side. To identify a sequence on the beta(3) subunit important for assembly, we generated various full-length or truncated chimeric beta(3) constructs and investigated their ability to assemble with alpha(1) and gamma(2) subunits. It was demonstrated that replacement of the sequence beta(3) (76-89) by the homologous alpha(1) sequence impaired assembly with alpha(1) but not with gamma(2) subunits in alpha(1) beta(3) gamma(2) -GABA(A) receptors. Other experiments indicated that assembly was impaired via the beta(3) (-) side of the chimeric subunit. Within the sequence beta(3) (76-89) the sequence beta(3) (85-89) seemed to be of primary importance for assembly with alpha(1) subunits. A comparison with the structure of the acetylcholine-binding protein supports the conclusion that the sequence beta(3) (85-89) is located at the beta(3) (-) side and indicates that it contains amino acid residues that might directly interact with the (+) side of the neighbouring alpha(1) subunit.