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|Autori: ||Magrì, Andrea|
|Titolo: ||VDAC and SOD1: two major players in mitochondrial metabolism and in ALS|
|Abstract: ||The Thesis work presented here has been devoted to two proteins, the Voltage-Dependent Anion Channel (VDAC) and the Superoxide Dismutase I (SOD1) and has been especially focused on the relationships between them in physiological or pathological conditions of the cell.
VDAC is a pore-forming protein located in the outer mitochondrial membrane, where it is suspected to play a key role in metabolism regulation, as the interface between mitochondria and cytosol, and in apoptosis regulation. In the small family of VDAC proteins, composed of three isoforms in chordates, VDAC3 is the least known. Conversely from isoforms 1 and 2, its ability to form pores has been questioned. In this thesis, we present the first complete electrophysiological characterization of VDAC3, showing that this protein is able to forms smaller pores compared to VDAC1, under physiological condition of pH. Another point examined here has been the gating of VDAC1. This protein, in vitro, shows the important feature of gating the pore in dependence of high voltage applied. It is believed that the N-terminal domain has a crucial role in voltage-dependent gating and in the stabilization of the protein through its interaction with the pore-wall.
By producing several mutants of VDAC1, in this work we have shown that the Voltage-dependence may be modulated in an asymmetrical way by modifying sequences or deleting £]-strands required for the interaction with N-terminal domain.
The Superoxide Dismutase I (SOD1) is the most important antioxidant enzyme of all eukaryotic cells, since it inactivates the superoxide anion. Many recent evidences suggest that SOD1 is important for mitochondrial function, both in physiological and pathological conditions. SOD1 protects, among others, VDAC from oxidative stress, and may affect mitochondrial proteins expression levels. In addition, in the neurodegenerative disease Amyotrophic Lateral Sclerosis (ALS), SOD1 mutants were reported to directly bind the cytosolic surface of mitochondria, using VDAC1 as docking site. To understand the relationships between VDAC and SOD1, in this work, we studied the influence of hSOD1 on mitochondria when it was overexpressed in a yeast strain devoid of yeast endogenous VDAC. Our results sturprisingly indicate that SOD1 may have a metabolic role and can support the mitochondrial recovery in the strain ´por1, heavily slowed down by the lack of porin1. Our results support the recent claim that SOD1 may act on the expression of other mitochondrial proteins. In addition, the characterization of the interaction between VDAC1 and two of the most diffused ALS-linked SOD1 mutants was also obtained and might be considered the molecular basis in understanding the mitochondrial involvement in ALS.|
|In||Area 05 - Scienze biologiche|
|MGRNDR84E01C351L-Tesi definitiva AM.pdf||Tesi Andrea Magrì||5,15 MB||Adobe PDF||Visualizza/apri
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