Biochemical and structural studies of yeast Vps4 oligomerization.
Biochemical and structural studies of yeast Vps4 oligomerization.
复制标题
DOI：
10.1016/j.jmb.2008.09.066
复制
发表时间：
2008-12-26
影响因子：
5.6
通讯作者：
Hill, Christopher P.
中科院分区：
文献类型：
--
作者：
Gonciarz, Malgorzata D.;Whitby, Frank G.;Eckert, Debra M.;Kieffer, Collin;Heroux, Annie;Sundquist, Wesley I.;Hill, Christopher P.
ESCRT途径在多囊泡体的囊泡形成、被包裹的RNA病毒(如HIV-1)的萌发以及细胞质分裂的最终脱落阶段发挥作用。作为ESCRT途径中唯一已知的酶，AAA ATPase Vps4提供了多轮囊泡形成所需的能量。像其他Vps4蛋白一样，酵母Vps4通过两种状态循环：一种是催化不活跃的分解状态，我们在这里展示的是一种二聚体；另一种是催化活性的高阶组装，我们已经建模为由两个堆叠的六聚体环组成的十二聚体。我们还报道了酵母VPS4蛋白在脱脂蛋白和三磷酸腺苷γS结合状态下的晶体结构。在这两种情况下，Vps4亚基组装成连续的螺旋，具有六重螺旋轴，类似于以前在其他Vps4晶体形式中看到的螺旋。通过相邻Vps4亚基的大小AAA ATPase结构域之间的广泛相互作用来稳定螺旋，这表明这些接触面可以用来构建催化活性的十二聚体和催化不活性的二聚体。与这一模型一致，我们已经确定了专门抑制Vps4二聚化和/或十二聚体的界面突变体。因此，Vps4二聚体和十二聚体可能形成截然不同但重叠的界面。最后，我们的结构研究使我们能够对一个保守的环(孔环2)的构象进行建模，该环被预测在Vps4六面环之一的中心形成一个富含精氨酸的孔。我们的突变分析表明，孔环2残基Arg241和Arg251是HIV-1有效萌发所必需的，从而支持这个“精氨酸环”在Vps4功能中的作用。
The ESCRT pathway functions in vesicle formation at the multivesicular body, the budding of enveloped RNA viruses such as HIV-1, and the final abscission stage of cytokinesis. As the only known enzyme in the ESCRT pathway, the AAA ATPase Vps4 provides the energy required for multiple rounds of vesicle formation. Like other Vps4 proteins, yeast Vps4 cycles through two states: a catalytically inactive disassembled state that we show here is a dimer, and a catalytically active higher order assembly that we have modeled as a dodecamer composed of two stacked hexameric rings. We also report crystal structures of yeast Vps4 proteins in the apo- and ATPγS-bound states. In both cases, Vps4 subunits assembled into continuous helices with six-fold screw axes that are analogous to helices seen previously in other Vps4 crystal forms. The helices are stabilized by extensive interactions between the large and small AAA ATPase domains of adjacent Vps4 subunits, suggesting that these contact surfaces may be used to build both the catalytically active dodecamer and catalytically inactive dimer. Consistent with this model, we have identified interface mutants that specifically inhibit Vps4 dimerization, dodecamerization, or both. Thus, the Vps4 dimer and dodecamer likely form distinct but overlapping interfaces. Finally, our structural studies have allowed us to model the conformation of a conserved loop (Pore Loop 2) that is predicted to form an arginine-rich pore at the center of one of the Vps4 hexameric rings. Our mutational analyses demonstrate that Pore Loop 2 residues Arg241 and Arg251 are required for efficient HIV-1 budding, thereby supporting a role for this “arginine collar” in Vps4 function.
登录
查看更多内容
DOI：
10.1083/jcb.200508166
发表时间：
2006-02-27
期刊：
The Journal of cell biology
影响因子：
--
作者：
Azmi I;Davies B;Dimaano C;Payne J;Eckert D;Babst M;Katzmann DJ
通讯作者：
Katzmann DJ
影响因子：
3.3
作者：
Bishop, N;Woodmane, P
通讯作者：
Woodmane, P
DOI：
10.1107/s0907444904019158
发表时间：
2004-12-01
影响因子：
2.2
作者：
Emsley, P;Cowtan, K
通讯作者：
Cowtan, K
DOI：
10.1038/nsb972
发表时间：
2003-10-01
期刊：
NATURE STRUCTURAL BIOLOGY
影响因子：
--
作者：
DeLaBarre, B;Brunger, AT
通讯作者：
Brunger, AT
影响因子：
56.9
作者：
Carlton, Jez G.;Martin-Serrano, Juan
通讯作者：
Martin-Serrano, Juan