Multiple structural elements determine subunit specificity of Mg2+ block in NMDA receptor channels.

In NMDA receptor channels, subtype-specific differences of Mg2+ block are determined by the NR2 subunits. Channels assembled from the NR1-NR2A or NR1-NR2B subunits are blocked more strongly than channels formed by the NR1-NR2C or NR1-NR2D subunits, predominantly reflecting a difference in voltage dependence. A determinant of Mg2+ block common to the NR2 subunits is located in the M2 domain (N-site or Q/R/N-site). However, subunit-specific differences of block suggested that additional structural elements exist. Chimeric NR2 subunits were constructed by replacing segments of the least sensitive NR2C subunit with homologous segments of the most sensitive NR2B subunit. Mutant NR2 subunits were coexpressed with wild-type NR1 in Xenopus oocytes, and Mg2+ block was quantified. Replacement of the entire M1-M4 region resulted in a chimera with a sensitivity of Mg2+ block similar to that of the NR2B wild type. Replacing smaller segments or introducing point mutations did not generate channels with Mg2+ block characteristic of NR2B wild type. However, combining in a single chimera three small segments (M1, M2-M3 linker, M4), each independently mediating an increase in Mg2+ block, produced channels close to NR2B wild type. Thus, differences in Mg2+ block as controlled by the NR2 subunits cannot be explained by a single structural determinant in addition to the N-site. Moreover, three elements of the NR2 subunit are the major determinants of subtype-specific differences of Mg2+ block in heteromeric NMDA receptor channels.