Mechanistic characterizations of redox regulations and functions of Arf and Rho families

项目总结:

Project Summary: Small GTPases play critical roles in cellular functions. They can be classified into several groups, including the Arf, Rab, Ras, Rho, and Ran families of proteins. The redox regulations and functions of the Arf families of small GTPases are unknown. The redox regulations and functions of the Rho families of small GTPases are only partly known. Without knowing them, the development of effective therapeutics for diseases associated with them is impractical. One long-term objective of this project is to understand the redox-dependent regulations and functions of these proteins. Another objective is to develop therapeutic interventions for these diseases. This objective includes the identification of the novel redox inert nucleotides that seem to target the redox- sensitive Arf and Rho proteins to block their redox responses. To achieve these goals, the previously unknown redox-sensitive motifs found in the Arf families of proteins will be characterized by using a multidisciplinary approach that includes mutagenesis-based redox biochemistry with novel mechanism-based nucleotide analogs as well as mass spectrometric, EPR spectroscopic, and cell biology methods. The multidisciplinary approach will also be used to refine and characterize previously unknown redox-sensitive motifs found in Rho proteins. The characterizations gained by using mutagenesis-based redox biochemistry and cell biology as well as other analytical methods will identify the unprecedented regulatory features and redox response properties of the new and refined redox-sensitive motifs found in Arf and Rho GTPases. The feature characteristics of these redox motifs in these proteins will then be classified based on their novel redox response properties. These classifications will further allow a systematic mechanistic investigation of the detailed redox control and action of these redox motifs in these small GTPases. This systematic mechanistic study will then specify the regulations of and mechanisms of the actions of the new and refined redox-sensitive motifs found in the Arf and Rho families of small GTPases. Furthermore, the redox biochemical analysis of this multidisciplinary approach includes the use of nucleotide analogs that are novel redox-inert nucleotides. The use of these analogs is not only to inspect the mechanisms of the actions of these redox motifs in the small GTPases but also to provide insight into a novel therapeutic strategy in which a redox desensitization of these motifs alleviates or terminates diseases associated with the unchecked redox response of these motifs in the small GTPases.