Crystal structure of the Sec4p-Sec2p complex in the nucleotide exchanging intermediate state
Crystal structure of the Sec4p-Sec2p complex in the nucleotide exchanging intermediate state
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DOI：
10.1073/pnas.0701550104
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发表时间：
2007-05-15
影响因子：
11.1
通讯作者：
Nureki, Osamu
中科院分区：
文献类型：
--
作者：
Sato, Yusuke;Fukai, Shuya;Nureki, Osamu
胞吐过程中的囊泡转运受Rab GT3（酵母中的Sec 4p）调节，其由称为Sec 2 p的鸟嘌呤核苷酸交换因子（GEF）激活。在这里，我们以2.7 A的分辨率报告了具有无核苷酸Sec 4p的复合物中Sec 2 p GEF结构域的晶体结构。在复合物形成时，Sec 2 p螺旋彼此接近，将Phe-109的侧链翻转到Sec 2 p的Leu-104和Leu-108。这三个残基提供疏水平台以吸引Sec 4p的开关I区中的Phe-49、Ile-53和Ile-55的侧链以及开关间区中的Phe-57和Trp-74。因此，开关I和II区域在很大程度上变形，以产生紧密配合Sec 2 p卷曲螺旋表面的平坦疏水界面。这些剧烈的构象变化破坏了开关I和结合的鸟嘌呤核苷酸之间的相互作用，这有利于GDP的释放。与最近报道的Sec4p.Sec2p复合物的3.3埃结构不同，我们的结构包含与P-环结合的磷酸根离子，这可能代表核小体交换反应的中间状态。
Vesicular transport during exocytosis is regulated by Rab GTPase (Sec4p in yeast), which is activated by a guanine nucleotide exchange factor (GEF) called Sec2p. Here, we report the crystal structure of the Sec2p GEF domain in a complex with the nucleotide-free Sec4p at 2.7 A resolution. Upon complex formation, the Sec2p helices approach each other, flipping the side chain of Phe-109 toward Leu-104 and Leu-108 of Sec2p. These three residues provide a hydrophobic platform to attract the side chains of Phe-49, Ile-53, and Ile-55 in the switch I region as well as Phe-57 and Trp-74 in the interswitch region of Sec4p. Consequently, the switch I and II regions are largely deformed, to create a flat hydrophobic interface that snugly fits the surface of the Sec2p coiled coil. These drastic conformational changes disrupt the interactions between switch I and the bound guanine nucleotide, which facilitates the GDP release. Unlike the recently reported 3.3 angstrom structure of the Sec4p.Sec2p complex, our structure contains a phosphate ion bound to the P-loop, which may represent an intermediate state of the nucleoticle exchange reaction.