Laboratory of Molecular Biology, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, US Department of Health and Human Services, Bethesda, MD, USA.
EMBO J. 2010 Mar 17;29(6):1045-54. doi: 10.1038/emboj.2010.6. Epub 2010 Feb 11.
VHS (Vps27, Hrs, and STAM) domains occur in ESCRT-0 subunits Hrs and STAM, GGA adapters, and other trafficking proteins. The structure of the STAM VHS domain-ubiquitin complex was solved at 2.6 A resolution, revealing that determinants for ubiquitin recognition are conserved in nearly all VHS domains. VHS domains from all classes of VHS-domain containing proteins in yeast and humans, including both subunits of ESCRT-0, bound ubiquitin in vitro. ESCRTs have been implicated in the sorting of Lys63-linked polyubiquitinated cargo. Intact human ESCRT-0 binds Lys63-linked tetraubiquitin 50-fold more tightly than monoubiquitin, though only 2-fold more tightly than Lys48-linked tetraubiquitin. The gain in affinity is attributed to the cooperation of flexibly connected VHS and UIM motifs of ESCRT-0 in avid binding to the polyubiquitin chain. Mutational analysis of all the five ubiquitin-binding sites in yeast ESCRT-0 shows that cooperation between them is required for the sorting of the Lys63-linked polyubiquitinated cargo Cps1 to the vacuole.
VHS（Vps27、Hrs 和 STAM）结构域存在于 ESCRT-0 亚基 Hrs 和 STAM、GGA 衔接蛋白和其他运输蛋白中。STAM VHS 结构域-泛素复合物的结构在 2.6Å 分辨率下得到解决，揭示了几乎所有 VHS 结构域中都保守的泛素识别决定因素。来自酵母和人类中所有包含 VHS 结构域的蛋白类别的 VHS 结构域，包括 ESCRT-0 的两个亚基，在体外都能结合泛素。ESCRTs 被认为参与了 Lys63 连接的多泛素化货物的分拣。完整的人源 ESCRT-0 结合 Lys63 连接的四聚泛素的亲和力比单泛素高 50 倍，尽管比 Lys48 连接的四聚泛素高 2 倍。这种亲和力的增加归因于 ESCRT-0 中灵活连接的 VHS 和 UIM 基序在与多泛素链的强烈结合中的协同作用。酵母 ESCRT-0 中所有五个泛素结合位点的突变分析表明，它们之间的合作对于 Lys63 连接的多泛素化货物 Cps1 到液泡的分拣是必需的。
