The Proton Responsiveness in the Extracellular Domain of GLIC Differs in the Presence of the ELIC Transmembrane Domain
The Proton Responsiveness in the Extracellular Domain of GLIC Differs in the Presence of the ELIC Transmembrane Domain
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DOI：
10.1021/acs.biochem.6b00900
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发表时间：
2017-04-18
期刊：
影响因子：
2.9
通讯作者：
Lummis, Sarah C. R.
中科院分区：
文献类型：
--
作者：
Alqazzaz, Mona A.;Price, Kerry L.;Lummis, Sarah C. R.
Cys环受体的脯氨酸同源物已被证明可用于理解其真核对应物，但即使是这些中研究最好的Gloeetamine配体门控离子通道（GLIC），仍然尚未完全理解。GLIC在pH（50）为5 - 6时被质子激活，表明组氨酸残基参与了其激活，但尽管已知成孔α-螺旋（M2）中的组氨酸残基（His 11 '）参与门控，但胞外结构域（ECD）中的His（His 127）不参与门控。然而，有证据表明GLIC甘氨酸嵌合体在GLIC胞外结构域中具有质子敏感残基或区域。在此，我们创建了具有GLIC的ECD和ELIC的跨膜结构域（GELIC）的新型嵌合受体。该受体在卵母细胞中的表达揭示了质子活化，尽管pH（50）（6.7）不同于GLIC（5.4）。对ECD中可质子化残基的探索揭示了5个Asp残基（31、49、91、136和178）的pK（a）在GLIC的开放和闭合状态之间不同。用Ala或Asn取代这些残基对Asp 91突变体中的GLIC和GELIC显示出某种程度上相似的效果，但对其他突变体显示出不同的效果。总体而言，数据表明ECD中残基的质子化是GELIC中通道开放的要求，但在GLIC中仅起次要作用，其中门控可能主要通过其孔中His残基的质子化来驱动。
Prokaryotic homologues of Cys-loop receptors have proven to be useful in understanding their eukaryotic counterparts, but even the best studied of these, Gloeobacter ligand-gated ion channel (GLIC), is still not yet fully understood. GLIC is activated by protons with a pH(50) between 5 and 6, implicating a histidine residue in its activation, but although a histidine residue (His11') in the pore-forming alpha-helix (M2) is known to be involved in gating, the His in the extracellular domain (ECD), His127, is not. Nevertheless, there is evidence from a GLIC glycine chimera for a proton sensitive residue or region in the GLIC extracellular domain Here we create a novel chimeric receptor with the ECD of GLIC and the transmembrane domain of ELIC (GELIC). Expression of this receptor in oocytes reveals proton activation, although the pH(50) (6.7) differs from that of GLIC (5.4). Exploration of protonatable residues in the ECD reveals that the pK(a)s of five Asp residues (31, 49, 91, 136, and 178) differ between the open and closed states of GLIC. Substitution of these residues with Ala or Asn shows somewhat similar effects for GLIC and GELIC in Asp91 mutants, but different effects for the others. Overall, the data suggest that protonation of residues in the ECD is a requirement for channel opening in GELIC but plays only a minor role in GLIC, where gating may be largely driven via protonation of the His residue in its pore.