Use of concatamers to study GABAA receptor architecture and function: application to delta-subunit-containing receptors and possible pitfalls

Sigel, Erwin; Kaur, Kuldeep H; Lüscher, Benjamin P; Baur, Roland (2009). Use of concatamers to study GABAA receptor architecture and function: application to delta-subunit-containing receptors and possible pitfalls. Biochemical Society transactions, 37(Pt 6), pp. 1338-42. London: Portland Press 10.1042/BST0371338

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Many membrane proteins, including the GABA(A) [GABA (gamma-aminobutyric acid) type A] receptors, are oligomers often built from different subunits. As an example, the major adult isoform of the GABA(A) receptor is a pentamer built from three different subunits. Theoretically, co-expression of three subunits may result in many different receptor pentamers. Subunit concatenation allows us to pre-define the relative arrangement of the subunits. This method may thus be used to study receptor architecture, but also the nature of binding sites. Indeed, it made possible the discovery of a novel benzodiazepine site. We use here subunit concatenation to study delta-subunit-containing GABA(A) receptors. We provide evidence for the formation of different functional subunit arrangements in recombinant alpha(1)beta(3)delta and alpha(6)beta(3)delta receptors. As with all valuable techniques, subunit concatenation has also some pitfalls. Most of these can be avoided by carefully titrating and minimizing the length of the linker sequences joining the two linked subunits and avoiding inclusion of the signal sequence of all but the N-terminal subunit of a multi-subunit construct. Maybe the most common error found in the literature is that low expression can be overcome by simply overloading the expression system with genetic information. As some concatenated constructs result by themselves in a low level of expression, this erroneous assembly leading to receptor function may be promoted by overloading the expression system and leads to wrong conclusions.

Item Type:	Journal Article (Original Article)
Division/Institute:	04 Faculty of Medicine > Pre-clinic Human Medicine > Institute of Biochemistry and Molecular Medicine
UniBE Contributor:	Sigel, Erwin, Lüscher, Benjamin, Baur, Roland
ISSN:	0300-5127
Publisher:	Portland Press
Language:	English
Submitter:	Factscience Import
Date Deposited:	04 Oct 2013 15:12
Last Modified:	05 Dec 2022 14:22
Publisher DOI:	10.1042/BST0371338
PubMed ID:	19909272
Web of Science ID:	000272646000035
URI:	https://boris.unibe.ch/id/eprint/31631 (FactScience: 196262)