RABBIT MUSCLE ALDOLASE

该子项目是利用NIH/NCRR资助的中心赠款提供的资源的许多研究子项目之一。子项目和研究者（PI）可能从另一个NIH来源获得主要资金，因此可以在其他CRISP条目中表示。所列机构为中心，不一定是研究者所在机构。醛缩酶是从原核生物到哺乳动物的所有生物中普遍存在的酶。它们具有裂解碳-碳键的能力。它们的作用在糖酵解中最为人所知，其中果糖-1，6-二磷酸（FBP）醛缩酶将FBP切割成二羟丙酮-磷酸（DHAP）和甘油醛-3-磷酸。由于缺乏真正的反应中间体晶体结构，残基的催化作用受到阻碍。 本课题的目的是研究兔肌醛缩酶的催化机制，该酶是一种Ⅰ类果糖-1，6-二磷酸醛缩酶。我们正计划使用本地和突变型醛缩酶晶体的底物浸泡实验，以了解催化中涉及的活性位点的残基。通过使用不同浸泡时间的组合，我们希望在反应方案中捕获催化反应中间体，已知其涉及3种共价中间体的形成。 时间分辨晶体学在兔肌肉醛缩酶晶体中的应用已经取得了成功。该技术为进一步研究醛缩酶的催化机理提供了可能。

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. Aldolase are ubiquitous enzymes found in all organisms, from prokaryotes to mammals. They have the ability to cleave carbon-carbon bonds. Their role is best known in glycolysis where fructose-1,6-bisphosphate (FBP) aldolase cleaves FBP to dihydroxyacetone-phosphate (DHAP) and glyceraldehy-3-phosphate. The catalytic role assigment to residues has beeen hampered by the lack of genuine reaction intermediate crystallographic structures. The aim of this project is to study the catalytic mechanism of the rabbit muscle aldolase, which is a class I fructose-1,6-bisphosphate aldolase. We are planning to use subtrate soaking experiments with native and mutant aldolase crystals to get insight about residues of the active site implicated in the catalysis. By using a combination of diverse soaking time, we want to trap catalytic reaction intermdiates among the reaction scheme, which is known to implicate the formation of 3 covalent intermediates. Time-resolved crystallography applied to rabbit muscle aldolase crystal soaked with ligands or substrates have been successfully used in the past years. Further insight into the catalytic mechanism of aldolase still possible by using this technique.