STRUCTURE OF THE HUMAN HUWE1 HECT DOMAIN

这个子项目是利用资源的许多研究子项目之一。 由NIH/NCRR资助的中心拨款提供。对子项目的主要支持 子项目的首席调查员可能是由其他来源提供的， 包括美国国立卫生研究院的其他来源。为子项目列出的总成本可能 表示该子项目使用的中心基础设施的估计数量， 不是由NCRR赠款提供给次级项目或次级项目工作人员的直接资金。 E3泛素连接酶催化泛素结合的最后一步，调节许多细胞过程。Hect类E3泛素(Ub)连接酶直接将Ub从结合的E2酶转移到多种底物上。Hect Ub连接酶的催化结构域具有分离E2结合区和催化部位的双叶结构。关于Hect结构域功能的一个重要问题是连接酶活性和特异性的控制。在这里，我们基于晶体结构对E3连接酶HUWE1的Hect结构域进行了功能分析，并表明单个N-末端螺旋显著稳定了Hect结构域。我们观察到，该元件调节Hect结构域的活性，通过在没有这个螺旋的情况下诱导的自我泛素化来测量，与它对底物Mcl-1的Ub结合的影响不同。这种对蛋白质的细微变化可能是Hect结构域E3连接所显示的大量底物特异性的核心。在X射线中获得了假想的蛋白质PODANSg2158的衍射性晶体。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. E3 ubiquitin ligases catalyze the final step of ubiquitin conjugation and regulate numerous cellular processes. The HECT class of E3 ubiquitin (Ub) ligases directly transfers Ub from bound E2 enzyme to a myriad of substrates. The catalytic domain of HECT Ub ligases has a bilobal architecture that separates the E2 binding region and catalytic site. An important question regarding HECT domain function is the control of ligase activity and specificity. Here we present a functional analysis of the HECT domain of the E3 ligase HUWE1 based on crystal structures and show that a single N-terminal helix significantly stabilizes the HECT domain. We observe that this element modulates HECT domain activity, as measured by self-ubiquitination induced in the absence of this helix, as distinct from its effects on Ub conjugation of substrate Mcl-1. Such subtle changes to the protein may be at the heart of the vast spectrum of substrate specificities displayed by HECT domain E3 ligases.Diffraction quality crystals of the hypothetical protein PODANSg2158 were obtained in the X-ray.