DNA Accelerated Oxidized SOD1 Aggregation And It's Cytotoxicity To HepG2 Cell Under Physiological And Ascorbate Condition

Amyotrophic lateral sclerosis (ALS) is one of the most common adult neurodegenerative diseases with unknown causes. Although it is well established that SOD1 mutants involves development of ALS, and causes motor neuron death through an as-yet unidentified gain of one or more injurious properties, little is currently known about the roles of SOD1 in neurocytotoxicity. DNA is the source of genetic information and possesses many important roles in life process. Even the weak damage in DNA could have profound effect on viability and genetic stability. Moreover, DNA damage in human cells and organs has been found to be associated with neurodegenerative disease. Here, the research on the interaction between SOD1 and DNA has been carried out. The main results are as follows:1 The endogenous fluorescence of SOD1 was quenched as the concentration and reaction time of ascorbate increasing .Exogenous fluorescence dyeing experiments showed that the alteration in hydrophobicity of SOD1 had a positive correlation with the concentration and reaction time of ascorbate.2 The aggregation behavior of SOD1 in the presence of DNA and ascorbate were examined under neutral conditions , which could mimic the effect of mutations and reflect the practical process done under physiological conditions to a high extent. GeenGreen binding experiment indicated that SOD1 and DNA probably binded together through electrostatic interaction. RALS (right angle light scattering) and DLS (dynamic light scattering) experiments indicated that DNA could act as a template to trigger the aggregation of SOD1 oxidized by ascorbate, the size of the aggregation had a positive correlation with the concentration of DNA,SOD1 and reaction time.Moreover the results revealed that a significant alteration in hydrophobicity of SOD1 caused by ascorbate, and the enrichment in SOD1 along DNA double strands were two main reasons responsible for DNA-accelerated SOD1 aggregation. 3 The morphology of DNA-accelerated SOD1 aggregates formed under neutral and ascorbate conditions was examined by transmission electron microscope. Several types of SOD1 aggregates were observed, which depended on the concentration and reaction time of ascorbate and the ratio of SOD1 to DNA. When the ration of SOD1 to DNA was fixed, the aggregations structure could be more compacter and steadier with longer reaction time.4 HepG2 cell was cultivated in the presence of DNA-accelerated SOD1 aggregations gained previously for a certain while, after this the viability was measured with MTT. The results revealed that ,under the setted conditions, with the ascorbste concentration and reaction time increasing the viability of HepG2 cell decreased, in another word the larger the size of the aggregation was, the higher the cytotoxicity of the aggregation was.Thus it could be seen there was a certain correlation between the cytotoxicity and the size of the aggragation...