Sequence and expression of a frog brain complementary DNA encoding a kainate-binding protein.
Excitatory amino acids (EAAs) are important neurotransmitters in the vertebrate central nervous system. Electrophysiological and ligand-binding studies indicate that at least three different receptor subtypes for EAAs exist--N-methyl-D-aspartate, kainate and quisqualate receptor subtypes--on the basis of the preferred agonist of the receptors. We recently purified a kainate-binding protein (KBP) from frog (Rana pipiens berlandieri) brain by domoic acid (a high-affinity kainate analogue) affinity chromatography, and showed that the kainate-binding activity was associated with a protein of relative molecular mass 48,000 (Mr 48 K). The pharmacological properties and the anatomical distribution of KBP were consistent with those of a kainate receptor-ionophore complex. We have now isolated a complementary DNA encoding KBP of Mr 48 K. The deduced amino-acid sequence of the KBP has similar hydrophobic profiles to those found in other ligand-gated ion channel subunits, and shows some amino-acid sequence similarities to the corresponding regions of brain nicotinic acetylcholine receptor subunits. Localization of the KBP messenger RNAs by in situ hybridization histochemistry is compatible with the results of immunohistochemistry and receptor autoradiography studies. COS-7 cells transfected with the cDNA encoding the KBP show high-affinity kainate-binding activity with pharmacological properties similar to those of the biochemically purified KBP. These results provide the first molecular characterization of an EAA-binding site and raise the possibility that the KBP cDNA encodes a ligand-binding subunit of a kainate receptor-ionophore complex.
Wada
K
K
Laboratory of Molecular Otology, NIDCD, Bethesda, Maryland 20892.
Dechesne
C J
CJ
Shimasaki
S
S
King
R G
RG
Kusano
K
K
Buonanno
A
A
Hampson
D R
DR
Banner
C
C
Wenthold
R J
RJ
Nakatani
Y
Y
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001923:Brain Chemistry; D002460:Cell Line; D004247:DNA / Q000235:genetics* / Q000302:isolation & purification; D015870:Gene Expression; D006651:Histocytochemistry; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009693:Nucleic Acid Hybridization; D012333:RNA, Messenger / Q000032:analysis / Q000235:genetics; D011894:Rana pipiens; D018092:Kainic Acid Receptors; D017981:Receptors, Neurotransmitter / Q000235:genetics*; D011978:Receptors, Nicotinic; D012689:Sequence Homology, Nucleic Acid; D014162:Transfection
D004247:DNA; D012333:RNA, Messenger; D018092:Kainic Acid Receptors; D017981:Receptors, Neurotransmitter; D011978:Receptors, Nicotinic