A single amino acid determines the subunit-specific spider toxin block of alpha-amino-3-hydroxy-5-methylisoxazole-4-propionate/kainate receptor channels.
Joro spider toxin (JSTX) is one of the most potent antagonists of glutamatergic AMPA/KA (alpha-amino-3-hydroxy-5-methylisoxazole-4-propionate/kainate) receptor channels in invertebrates and vertebrates. A differential blocking effect on certain types of glutamatergic synapses--e.g., parallel and climbing fiber synaptic inputs to rat cerebellar Purkinje neurons--has been shown by using a synthetic analog of the spider toxin. By investigating the molecular basis of the JSTX action on the recombinant AMPA/KA receptors GluR1-GluR4 and GluR6 expressed in Xenopus oocytes, we found that submicromolar concentrations of JSTX exert a subunit-specific block. Thus, receptor subunits forming a receptor channel with a linear current-voltage (I-V) relationship (GluR1/2, GluR2/3, and GluR6) were not affected, while receptor subunits with rectifying I-V relationships (GluR1, GluR3, GluR4, and GluR1/3) were reversibly blocked by JSTX. By using receptor-subunit mutants obtained by site-directed mutagenesis, we have identified a single amino acid position (glutamine in the proposed second transmembrane domain) that is critical for the JSTX block. Since this site has previously been shown to control the I-V relationship of the AMPA/KA receptor channel and to participate in the regulation of the channel's permeability for calcium ions, our findings suggest that JSTX binds close to the central pore region of the channel.
Blaschke
M
M
Max-Planck-Institut für Biophysikalische Chemie, Göttingen, Germany.
Keller
B U
BU
Rivosecchi
R
R
Hollmann
M
M
Heinemann
S
S
Konnerth
A
A
D000595:Amino Acid Sequence; D000818:Animals; D001120:Arginine / Q000235:genetics; D002118:Calcium / Q000378:metabolism; D003217:Conductometry; D005973:Glutamine / Q000235:genetics; D008969:Molecular Sequence Data; D009498:Neurotoxins / Q000235:genetics* / Q000494:pharmacology; D009865:Oocytes; D014176:Protein Biosynthesis; D018091:Receptors, AMPA; D017470:Receptors, Glutamate / Q000096:biosynthesis / Q000187:drug effects*; D018092:Kainic Acid Receptors; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000187:drug effects; D017386:Sequence Homology, Amino Acid; D013111:Spider Venoms / Q000235:genetics* / Q000494:pharmacology; D013329:Structure-Activity Relationship; D014981:Xenopus
D001120:Arginine; D002118:Calcium; D005973:Glutamine; D009498:Neurotoxins; D018091:Receptors, AMPA; D017470:Receptors, Glutamate; D018092:Kainic Acid Receptors; D011994:Recombinant Proteins; D013111:Spider Venoms; C044936:JSTX spider toxin