Differential contribution of the NR1‐ and NR2A‐subunits to the selectivity filter of recombinant NMDA receptor channels.
Differential contribution of the NR1‐ and NR2A‐subunits to the selectivity filter of recombinant NMDA receptor channels.
复制标题
NR1 和 NR2A 亚基对重组 NMDA 受体通道选择性过滤器的不同贡献。
DOI：
10.1113/jphysiol.1996.sp021257
复制
发表时间：
1996
期刊：
影响因子：
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通讯作者：
B. Sakmann
中科院分区：
文献类型：
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作者：
L. Wollmuth;T. Kuner;P. Seeburg;B. Sakmann
1.在非洲爪蟾卵母细胞中研究了由野生型和突变型NR 1-和NR 2A-亚基组成的重组N-甲基-D-天冬氨酸（NMDA）受体通道狭窄收缩的分子决定因素。2.不同大小的有机阳离子的相对渗透性被用作狭窄收缩的大小的指标。根据在双离子条件下测量的反转电位，以K+作为参比溶液，使用刘易斯方程计算渗透率比。3.对于野生型NMDA受体通道，五种有机阳离子显示出明显的逆转电位，其渗透率比（PX/PK）为：铵，1.28;甲基铵，0.48;二甲基铵（DMA），0.20;二乙基铵，0.07;和二甲基乙醇铵，0.02。4.在NR 1（位置598）或NR 2A（位置595）中的同源位置处，N-位点天冬酰胺（N）突变为谷氨酰胺（Q），相对于野生型通道，DMA的通透性增加大致相等。然而，对于较大尺寸的有机阳离子，NR 1（N598 Q）突变有更强的效果，增加其渗透性，而NR 2A（N595 Q）突变没有效果。有机阳离子渗透性的这些变化表明，NR 1（N598 Q）突变增加了孔径，而NR 2A（N595 Q）突变没有。5.其中NR 1 N-位点天冬酰胺被较小的甘氨酸（G）NR 1（N598 G）-NR 2A取代的通道显示出在任一亚基中检查的所有位点中孔径的最大增加。相比之下，在NR 2A亚基中，相同的N位点取代为甘氨酸对孔径仅产生很小的影响。6.对于NR 2A-亚基，当突变为更大或更小尺寸的氨基酸时，N-位点C-末端侧的天冬酰胺残基（位置596）对孔径产生较大的体积特异性影响。突变通道NR 1-NR 2A（N596 G）在NR 2A-亚基中检查的所有位点中孔径增加最大。相反，NR 1-亚基中同源位置的突变对孔径没有影响。7.野生型NMDA受体通道狭窄收缩的横截面直径估计为0.55 nm。NR 1（N598 G）-NR 2A和NR 1-NR 2A（N596 G）突变体通道的孔径分别增加至约0.75和0.67 nm。双突变NR 1（N598 G）-NR 2A（596 G）将孔径增加到约0.87 nm，基本上是单个突变产生的增加的总和。8.结论是NR 1-和NR 2A-亚基都有助于NMDA受体通道的狭窄收缩，天冬酰胺位于非同源位置。NMDA受体通道狭窄收缩的主要决定因素是NR 1 N-位点天冬酰胺和邻近NR 2A N-位点的天冬酰胺。
1. The molecular determinants for the narrow constriction of recombinant N‐methyl‐D‐aspartate (NMDA) receptor channels composed of wild‐type and mutant NR1‐ and NR2A‐subunits were studied in Xenopus oocytes. 2. The relative permeability of differently sized organic cations was used as an indicator of the size of the narrow constriction. From measured reversal potentials under bi‐ionic conditions with K+ as the reference solution, permeability ratios were calculated with the Lewis equation. 3. For wild‐type NMDA receptor channels, five organic cations showed clear reversal potentials, with permeability ratios (PX/PK): ammonium, 1.28; methylammonium, 0.48; dimethylammonium (DMA), 0.20; diethylammonium, 0.07; and dimethylethanol‐ammonium, 0.02. 4. Mutation of the N‐site asparagine (N) to glutamine (Q) at homologous positions in either NR1 (position 598) or NR2A (position 595) increased the permeability of DMA relative to wild‐type channels about equally. However, for larger sized organic cations, the NR1(N598Q) mutation had stronger effects on increasing their permeability whereas the NR2A(N595Q) mutation was without effect. These changes in organic cation permeability suggest that the NR1(N598Q) mutation increases the pore size while the NR2A(N595Q) mutation does not. 5. Channels in which the NR1 N‐site asparagine was replaced by the smaller glycine (G), NR1(N598G)‐NR2A, showed the largest increase in pore size of all sites examined in either subunit. In contrast, in the NR2A‐subunit the same N‐site substitution to glycine produced only small effects on pore size. 6. For the NR2A‐subunit, an asparagine residue (position 596) on the C‐terminal side of the N‐site, when mutated to larger or smaller sized amino acids, produced large, volume‐specific effects on pore size. The mutant channel NR1‐NR2A(N596G) had the largest increase in pore size of all sites examined in the NR2A‐subunit. In contrast, mutation of the homologous position in the NR1‐subunit had no effect on pore size. 7. The cross‐sectional diameter of the narrow constriction in wild‐type NMDA receptor channels was estimated to be 0.55 nm. The pore sizes of the NR1(N598G)‐NR2A and NR1‐NR2A(N596G) mutant channels increased to approximately 0.75 and 0.67 nm, respectively. The double mutation, NR1(N598G)‐NR2A(596G), increased the pore size to approximately 0.87 nm, essentially the sum of the increase produced by the individual mutations. 8. It is concluded that both the NR1‐ and NR2A‐subunits contribute to the narrow constriction of NMDA receptor channels with asparagines located at non‐homologous positions. The major determinants of the narrow constriction in NMDA receptor channels are the NR1 N‐site asparagine and an asparagine adjacent to the NR2A N‐site.
神经递质门控离子通道的渗透途径。
DOI：
10.1146/annurev.bb.21.060192.001411
发表时间：
1992
期刊：
Annual review of biophysics and biomolecular structure
影响因子：
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作者：
Lester,HA
通讯作者：
Lester,HA