The Study Of Molecular Mechanism Of Neuroglobin And Cytoglobin Against Oxidative Injury

Oxygen is the essential element for life. How the life body utilizes oxygen is an important scientific proposition for health. The discoveries of oxygen-binding globins, neuroglobin and cytoglobin, have brought more complexity to know the process of oxygen metabolism. Neuroglobin specifically expresses in nervous tissue, however, cytoglobin widely distributes nearly all types of tissues. For that both of them has the analogous structure basis of binding to heme and the same oxygen bonding properties as other members of oxygen-binding globins family , such as hemoglobin and myoglobin, neuroglobin and cytoglobin were considered to protect bodies against oxidative injury. Several function hypotheses about neuroglobin and cytoglobin were speculated. However, there were no conclusive evidences to support anyone of these function hypothesis. Because proteins play their roles often through the molecular network formed by interaction with other proteins, we wish to study the function mechanisms of neuroglobin and cytoglobin from proteins interacting with them.This dissertation was processed with two parts. In the first part, we focused on the protein interacting with neuroglobin, glutamyl-prolyl tRNA synthetase, which was screened by yeast two-hybrid system to study the physiology mechanism of neuroglobin under hypoxia. At first, we demonstrated with various biochemical evidences that neuroglobin could interact with the neuroglobin. On the structure and function of glutamyl-prolyl tRNA synthetase, we screened and identified several aminoacyl tRNA synthetases such as histidyl-tRNA synthetase, glutaminyl-tRNA synthetase, isoleucine-tRNA synthetase and coenzyme factor P43 sensitive to oxygen deprivation by CoCI₂. All these results refer us the groundwork to study the role of aminoacyl tRNA synthetase in hypoxia of nervous system and also help us to clarify the molecular mechanism of neuroglobin protecting neuron against oxidative injury.In the second part, we studied the physiology mechanism of cytoglobin under hypoxia induced by CoCI₂ based on the interacting protein, metallothionein II, which was screened by yeast two-hybrid system. At first the interaction existing between cytoglobin and metallothionein II was further identified by co-immunoprecipitation assay followed yeast back transfection assay and cellular fluorescence co-localization assay. Then according to the reported physiological function of metallothionein II .which can detoxified the heavy metals toxicity and directly cleared off the free radicals, we successfully built various oxidative injury cellular models and found that overexpressed cytoglobin and metallothionein II can protect PC12 cells against oxidative insult. Based on which, we hypothesized that cytoglobin might recruit metallothionein II to strengthen the ability of directly clearing excess free radicals and then widely play a role against oxidative injury.In summary, we found the potential mechanisms for the protective function of neuroglobin and cytoglobin, which are critically important for novel drugs development for neuroprotective usage in clinical treatment for stroke and other hypoxia and ischemia related diseases.