A regulatory domain (R1-R2) in the amino terminus of the N-methyl-D-aspartate receptor: Effects of spermine, protons, and ifenprodil, and structural similarity to bacterial leucine/isoleucine/valine binding protein

There are complex interactions between spermine, protons, and ifenprodil at N-methyl-D-aspartate receptors. Spermine stimulation may involve relief of proton inhibition, whereas ifenprodil inhibition may involve an increase i n proton inhibition. We studied mutations at acidic residues in the NR1 sub unit using voltage-clamp recording of NR1/NR2B receptors expressed in Xenop us oocytes. Mutations at residues near the site of the exon-5 insert, inclu ding E181 and E185, reduced spermine stimulation and proton inhibition. Mut ation NR1(D130N) reduced sensitivity to ifenprodil by more than 500-fold, b ut had little effect on sensitivity to spermine and pH. Mutations at six ot her residues in this region of the NR1 subunit reduced the potency and, in some cases, the maximum effect of ifenprodil. These mutants did not affect sensitivity to pH, glutamate, glycine, or other hallmark properties of N-me thyl-D-aspartate channels such as Mg2+ block and Ba2+ permeability. Residue s in this region presumably form part of the ifenprodil-binding site. To mo del this region of NR1 we compared the predicted secondary structure of NR1 (residues 19-400) with the known structures of 1,400 proteins. This region of NR1 is most similar to bacterial leucine/isoleucine/valine binding prot ein, a globular amino acid binding protein containing two robes, similar to the downstream S1-S2 region of glutamate receptors. We propose that the te rtiary structure of NR1(22-375) is similar to leucine/isoleucine/valine bin ding protein, containing two "regulatory" domains, which we term R1 and R2. This region, which contains the binding sites for spermine and ifenprodil, may influence the downstream S1 and S2 domains that constitute the glycine binding pocket.