Comparative structure of human neuronal alpha 2-alpha 7 and beta 2-beta 4 nicotinic acetylcholine receptor subunits and functional expression of the alpha 2, alpha 3, alpha 4, alpha 7, beta 2, and beta 4 subunits.
cDNA clones encoding human neuronal nicotinic acetylcholine receptor alpha 2, alpha 3, alpha 4, alpha 5, alpha 6, alpha 7, beta 2, beta 3, and beta 4 subunits were isolated from brainstem, hippocampus, prefrontal cortex, substantia nigra, thalamus, and IMR32 libraries. Human alpha 2 and alpha 6 and full-length beta 3 and beta 4 clones have not been previously reported. Deduced amino acid sequences of the alpha 2, alpha 6, beta 3, and beta 4 predicted mature peptides are 503 residues (56.9 kDa), 464 residues (53.7 kDa), 440 residues (50.8 kDa), and 477 residues (54.1 kDa), respectively. These sequences show 84 (alpha 2), 87 (alpha 6), 89 (beta 3), and 84% (beta 4) identity to the corresponding rat sequences. The amino termini of the human alpha 2 and beta 3 mature peptides contain 23 and six additional residues, respectively, compared to those of rat alpha 2 and beta 3. Recombinant receptors were expressed in Xenopus laevis oocytes injected with in vitro transcripts encoding either alpha 7 alone or alpha 2, alpha 3, or alpha 4 in pairwise combination with beta 2 or beta 4. Inward currents were elicited by the application of acetylcholine (1-100 microM) and other agonists; these responses were blocked 65-97% by application of 10 microM d-tubocurare, confirming functional expression of human nicotinic receptors.
Elliott
K J
KJ
SIBIA Neurosciences Inc., La Jolla, CA, USA.
Ellis
S B
SB
Berckhan
K J
KJ
Urrutia
A
A
Chavez-Noriega
L E
LE
Johnson
E C
EC
Veliçelebi
G
G
Harpold
M M
MM
D000109:Acetylcholine / Q000494:pharmacology; D000595:Amino Acid Sequence; D000818:Animals; D003001:Cloning, Molecular; D018076:DNA, Complementary; D005260:Female; D015723:Gene Library; D006801:Humans; D046911:Macromolecular Substances; D008564:Membrane Potentials / Q000187:drug effects; D008969:Molecular Sequence Data; D009474:Neurons / Q000378:metabolism*; D009865:Oocytes / Q000502:physiology; D051381:Rats; D011978:Receptors, Nicotinic / Q000096:biosynthesis* / Q000737:chemistry* / Q000502:physiology; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry; D017386:Sequence Homology, Amino Acid; D014982:Xenopus laevis
D018076:DNA, Complementary; D046911:Macromolecular Substances; D011978:Receptors, Nicotinic; D011994:Recombinant Proteins; D000109:Acetylcholine