STRUCTURAL CHANGES IN HUMAN SOD1 BY POST-TRANLATIONAL MODIFICATION

这个子项目是许多利用资源的研究子项目之一 由NIH/NCRR资助的中心拨款提供。子项目的主要支持 而子项目的主要调查员可能是由其他来源提供的， 包括其它NIH来源。 列出的子项目总成本可能 代表子项目使用的中心基础设施的估计数量， 而不是由NCRR赠款提供给子项目或子项目工作人员的直接资金。 肌萎缩侧索硬化症（ALS）是一种以运动神经元死亡为特征的神经退行性疾病。大约20%的ALS遗传病例是由编码Cu，Zn超氧化物歧化酶（SOD 1）的基因突变引起的。这些突变的一个后果是引起蛋白质的聚集。这些突变发生在这个153个氨基酸蛋白质的70多个不同位点，使得难以确定共同的毒性机制。SOD 1是一种同源二聚体酶，每个亚基结合两个金属离子，并在C57和C146之间含有分子内二硫键。由于所有这些翻译后修饰都是完整的，这种酶具有高度的热稳定性，熔点超过90 ℃。最近的生物物理分析表明，不成熟的形式的酶，缺乏金属离子，二硫键或两者兼而有之，可能更有可能聚集。二硫键还原的无金属形式的酶具有边际稳定性，并且酶的ALS变体甚至可以在生理温度下展开。为了确定SOD 1单体形式的结构差异，我们计划用小角X射线散射（SAXS）研究WT蛋白的稳定单体版本和选定的一组ALS变体之间的全局结构差异。这将包括对SOD 1的研究，包括翻译后修饰的多个阶段、氧化和还原条件下以及金属负载的各个阶段。 该结果有望对我们理解疾病病理的分子机制产生重大影响

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. Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease characterized by the death of motor neurons. Approximately 20% of all inherited cases of ALS are caused by mutations in the gene encoding for Cu, Zn superoxide dismutase (SOD1). One consequence of these mutations is to cause the aggregation of the protein. These mutations occur at over 70 different sites of this 153 amino acid protein, making it difficult to ascertain a common toxic mechanism. SOD1 is a homodimeric enzyme which binds two metal ions per subunit and contains an intramolecular disulfide-bond between C57 and C146. With all of these post-translational modifications intact, the enzyme is highly thermostable, with a melting point over 90 ¿C. Recent biophysical analyses have suggested that premature forms of the enzyme, lacking either the metal ions, the disulfide bond or both, may be more likely candidates for aggregation. The disulfide-reduced, metal-free forms of the enzyme have marginal stability, and ALS-variants of the enzyme may even be unfolded at physiological temperatures. In order to determine structural differences in the monomeric forms of SOD1, we plan to study global structural differences between stable monomeric versions of the WT protein and a selected set of ALS variants with small angle x-ray scattering (SAXS). This will include studies of SOD1, with multiple stages of posttranslational modification and under oxidized and reduced conditions and also under various stages of metal loading. The results are expected to have an significant impact on our understanding of the molecular mechanism of disease pathology