ULTRA-HIGH RESOLUTION STUDIES OF CHOLESTEROL OXIDASE INCUBATED WITH XENON: HIV

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。对黄酶胆固醇氧化酶（ChO）进行了超高分辨率结构研究。ChO是一种55 kDa的双功能黄酶，参与类固醇的氧化和异构化，类固醇核的C5位置含有一个3-羟基和一个双键。这种酶在临床上被用于测定血清胆固醇，最近，由于这种酶能改变膜结构，它被用作一种有效的杀幼虫剂。此外，该酶也被认为是马红球菌的毒力因子之一，马红球菌是一种与人类免疫缺陷病毒感染有关的人类机会性病原体。在人类中，马红球菌感染主要影响肺部，其临床病理特征与免疫功能低下患者的肺结核相似。胆固醇氧化酶作用于真核细胞膜，其膜损伤特性可能是导致红球菌感染的细胞毒性和巨噬细胞破坏的原因。鉴于该酶在发病机制中的突出作用，我们相信，清楚地了解底物结合特性以及催化机制可能有助于开发用于治疗红球菌感染患者的抑制剂。利用氙孵育研究了氧气进入活性部位的通道。初步研究的数据集扩展到0.85¿¿（单元尺寸：a=51.267, b=73.213¿¿c=63.070¿¿alpha=90.0¿¿beta=105.132¿¿gamma=90）。虽然氙被发现可以结合，但它并不在所谓的通道中。

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. Ultra-high resolution structural studies of the flavoenzyme cholesterol oxidase (ChO) was carried out. ChO is a 55 kDa bifunctional flavoenzyme involved in the oxidation and isomerization of steroids that contain a 3-beta-hydroxyl group and a double bond at the C5 position of the steroid nucleus. The enzyme has been used clinically for the determination of serum cholesterol and most recently, due to the enzyme¿¿s ability to alter membrane structure, it has found applications as a potent larvicide. Moreover, the enzyme has also been proposed to be one of the virulence factors in Rhodococcus equi, an opportunistic pathogen in humans, associated with human immunodeficiency virus infection. In humans, Rhodococcus equi infection mainly affects the lungs with clinical pathological characteristics similar to pulmonary tuberculosis in immunocompromised patients. Cholesterol oxidase acts on eukaryotic membranes and its membrane damaging properties may be responsible for the observed cytotoxicity and macrophage destruction that accompanies Rhodococcal infection. Given the enzyme¿¿s prominent role in pathogenesis, we believe that a clear understanding of the substrate binding properties as well as the catalytic mechanism may aid in the development of inhibitors used as drugs to treat patients suffering from Rhodococcal infection. The purported channel for oxygen entry to the active site was studied using xenon incubation. Preliminary studies resulted in data sets extending to 0.85 ¿¿(cell dimensions: a=51.267, b=73.213 ¿¿ c=63.070 ¿¿ alpha=90.0 ¿¿ beta=105.132 ¿¿ gamma=90). Although xenon was found to bind, it was not in the purported channel.