Construction of covalently coupled, concatameric dimers of 7TM receptors
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Construction of covalently coupled, concatameric dimers of 7TM receptors. / Terpager, Marie; Scholl, D Jason; Kubale, Valentina; Martini, Lene; Elling, Christian E; Schwartz, Thue W.
I: Journal of Receptors and Signal Transduction, Bind 29, Nr. 5, 2009, s. 235-45.Publikation: Bidrag til tidsskrift › Tidsskriftartikel › Forskning › fagfællebedømt
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TY - JOUR
T1 - Construction of covalently coupled, concatameric dimers of 7TM receptors
AU - Terpager, Marie
AU - Scholl, D Jason
AU - Kubale, Valentina
AU - Martini, Lene
AU - Elling, Christian E
AU - Schwartz, Thue W
N1 - Keywords: Animals; CHO Cells; Cell Membrane; Cricetinae; Cricetulus; Cyclic AMP; Dimerization; Dipeptides; Humans; Inositol Phosphates; Membrane Proteins; Peptide Fragments; Protein Binding; Receptors, Adrenergic, beta-2; Receptors, G-Protein-Coupled; Receptors, Neurokinin-1; Structure-Activity Relationship; Substance P
PY - 2009
Y1 - 2009
N2 - 7TM receptors are easily fused to proteins such as G proteins and arrestin but because of the fact that their terminals are found on each side of the membrane they cannot be joined directly in covalent dimers. Here, we use an artificial connector comprising a transmembrane helix composed of Leu-Ala repeats flanked by flexible spacers and positively charged residues to ensure correct inside-out orientation plus an extracellular HA-tag to construct covalently coupled dimers of 7TM receptors. Such 15 TM concatameric homo- and heterodimers of the beta(2)-adrenergic and the NK(1) receptors, which normally do not dimerize with each other, were expressed surprisingly well at the cell surface, where they bound ligands and activated signal transduction in a manner rather similar to the corresponding wild-type receptors. The concatameric heterodimers internalized upon stimulation with agonists for either of the protomers, which was not observed upon simple coexpression of the two receptors. It is concluded that covalently joined 7TM receptor dimers with surprisingly normal receptor properties can be constructed with use of an artificial transmembrane connector, which perhaps can be used to fuse other membrane proteins.
AB - 7TM receptors are easily fused to proteins such as G proteins and arrestin but because of the fact that their terminals are found on each side of the membrane they cannot be joined directly in covalent dimers. Here, we use an artificial connector comprising a transmembrane helix composed of Leu-Ala repeats flanked by flexible spacers and positively charged residues to ensure correct inside-out orientation plus an extracellular HA-tag to construct covalently coupled dimers of 7TM receptors. Such 15 TM concatameric homo- and heterodimers of the beta(2)-adrenergic and the NK(1) receptors, which normally do not dimerize with each other, were expressed surprisingly well at the cell surface, where they bound ligands and activated signal transduction in a manner rather similar to the corresponding wild-type receptors. The concatameric heterodimers internalized upon stimulation with agonists for either of the protomers, which was not observed upon simple coexpression of the two receptors. It is concluded that covalently joined 7TM receptor dimers with surprisingly normal receptor properties can be constructed with use of an artificial transmembrane connector, which perhaps can be used to fuse other membrane proteins.
U2 - 10.1080/10799890903154217
DO - 10.1080/10799890903154217
M3 - Journal article
C2 - 19747085
VL - 29
SP - 235
EP - 245
JO - Journal of Receptor and Signal Transduction Research
JF - Journal of Receptor and Signal Transduction Research
SN - 1079-9893
IS - 5
ER -
ID: 21666199