CATALYTIC MECHANISM AND INHIBITION OF GOLGI ALPHA-MANNOSIDASE II

本子项目是利用由NIH/NCRR资助的中心赠款提供的资源的众多研究子项目之一。子项目和研究者（PI）可能已经从另一个NIH来源获得了主要资金，因此可以在其他CRISP条目中表示。列出的机构是中心的，不一定是研究者的机构。我们之前确定了糖基化酶-甘露糖苷酶II的结构，这是一种被潜在的抗转移化合物靶向的38家族的糖基水解酶。主要利用CHESS的数据，我们已经通过中间体和几种抑制剂的配合物结构分析了酶的催化和抑制的原子基础。最近，我们对化学文库进行了两次高通量筛选，寻找新的酶抑制剂。这导致了10-12次不同亲和力和属性的命中。我们感兴趣的是研究它们是如何与结构结合的。特别是，有些可能是非竞争性抑制剂，并且可能占据远离催化中心的位点，这将是未来抑制剂设计的兴趣所在。我们想使用同步辐射有两个主要原因。首先，我们可以在几分钟内获得数据集，而不是几小时，这将使我们在短时间内快速检查这些复合体。其次，我们可以从这些晶体中获得几乎原子分辨率（1.2-1.4A）的数据。这种分辨率对于理解构象畸变和确定精确的原子间距离至关重要

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. We previously determined the structure of the glycosylation enzyme alpha-mannosidase II, a glycosyl hydrolase of the Family 38 family targetted by potential anti-metastatic compounds. Largely with data from CHESS, we have been analyzing the atomic basis of catalysis and inhibition of the enzyme through structures of complexes with intermediates and several classes of inhibitors. Recently, we performed two high-throughput screens of chemical libraries looking for novel inhibitors of the enzyme. This resulted in 10-12 hits of varying affinity and properties. We are interested in examining how these bind to the structure. In particular, some may be non-competitive inhibitors and may occupy sites distant from the catalytic centre, which will be of interest for future inhibitor design. We would like to use synchrotron radiation for two promary reasons. Firstly, we can get a data set in minutes, rather than hours, which will let us quickly examine several of these complexes in a short time. Secondly, we can get almost atomic resolution (1.2-1.4A) data from these crystals at CHESS. This resolution can be critical in understanding conformational distortions and determining precise interatomic dist