A short region upstream of the yeast vacuolar Qa-SNARE heptad-repeats promotes membrane fusion through enhanced SNARE complex assembly.
A short region upstream of the yeast vacuolar Qa-SNARE heptad-repeats promotes membrane fusion through enhanced SNARE complex assembly.
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DOI：
10.1091/mbc.e17-04-0218
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发表时间：
2017-08-15
影响因子：
3.3
通讯作者：
Wickner W
中科院分区：
文献类型：
--
作者：
Song H;Wickner W
膜融合需要四个SNARE结构域形成复合物。Qa-SNARE七重复结构域上游的一个短的保守区域促进SNARE复合物组装，从而促进融合。尽管SNARE（可溶性N-乙基马来酰亚胺敏感因子附着蛋白受体）七重重复序列已得到充分研究，但SNARE也具有不确定功能的上游N结构域。可以在化学确定的反应中研究酵母液泡SNARE组装成用于融合的复合物。将带有Rab：GTP和空泡R-SNARE或三个完整锚定的Q-SNARE之一的互补蛋白脂质体与系链/SM蛋白复合物HOPS和另外两个可溶性SNARE（缺乏跨膜锚）或其SNARE七肽重复结构域一起孵育。融合需要在一个膜上跨膜锚定的R-SNARE和在另一个膜上锚定的Q-SNARE。Qb-SNARE的N-结构域被完全包裹以用于融合。尽管融合可以通过非常高浓度的单独的Qa-SNARE七重重复结构域来促进，但是在生理浓度下，单独的Qa-SNARE七重重复结构域几乎没有融合活性。Qa的181-198区域，紧邻SNARE七重复结构域的上游，是与HOPS的正常融合活性所需的。该区域是正常SNARE复合体组装所需的。
Membrane fusion requires that four SNARE domains form a complex. A short conserved region just upstream of the Qa-SNARE heptad-repeat domain promotes SNARE-complex assembly and hence fusion. Whereas SNARE (soluble N-ethylmaleimide–sensitive factor attachment protein receptor) heptad-repeats are well studied, SNAREs also have upstream N-domains of indeterminate function. The assembly of yeast vacuolar SNAREs into complexes for fusion can be studied in chemically defined reactions. Complementary proteoliposomes bearing a Rab:GTP and either the vacuolar R-SNARE or one of the three integrally anchored Q-SNAREs were incubated with the tethering/SM protein complex HOPS and the two other soluble SNAREs (lacking a transmembrane anchor) or their SNARE heptad-repeat domains. Fusion required a transmembrane-anchored R-SNARE on one membrane and an anchored Q-SNARE on the other. The N-domain of the Qb-SNARE was completely dispensable for fusion. Whereas fusion can be promoted by very high concentrations of the Qa-SNARE heptad-repeat domain alone, at physiological concentrations the Qa-SNARE heptad-repeat domain alone has almost no fusion activity. The 181–198 region of Qa, immediately upstream of the SNARE heptad-repeat domain, is required for normal fusion activity with HOPS. This region is needed for normal SNARE complex assembly.