Structural and functional characterization of the disease-linked inner mitochondrial membrane protein MPV17

• Applicant: Professor Dr. Franz Hagn
• Subject Area: Structural Biology ; Biochemistry ; Biophysics
• Term: since 2024
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 540999464

The membrane protein MPV17 localized in the inner mitochondrial membrane is one of the causes of the so-called "mitochondrial DNA depletion" syndrome (MDDS), i.e. the reduction of the amount of DNA in the mitochondrial matrix. Since the essential components of the respiratory chain are encoded on mitochondrial DNA, mutations in the MPV17 gene lead to severe liver damage and hepatocerebral dysfunction, and thus usually to death in early infancy. The function and mechanistic details of MPV17 are relatively poorly understood, so that therapies are limited to the treatment of symptoms and are therefore not effective. In the project proposed here, we aim to better understand the molecular causes of MPV17-related diseases. To this end, we will first elucidate how MPV17 functions in order to clarify whether this protein can be a transporter of metabolites, e.g. precursors of nucleic acids. For this purpose, the direct binding of metabolites to MPV17, but also the effect of deletion of MPV17 on the mitochondrial metabolome will be investigated. A role of MPV17 as a voltage-gated proton channel discussed in the literature will also be investigated, which leads to a reduction of the membrane potential when the respiratory chain is overloaded, thus reducing the formation of reactive oxygen species (ROS). This can prevent damage to the cell, including the DNA in the mitochondria. Second, MPV17 has been associated with stabilization of mitochondrial substructures (cristae), which is directly linked to respiratory chain efficiency. To investigate this potential function, we will perform cryo-electron tomography studies using MPV17-knockout cells. With the high spatial resolution this allows, an accurate picture of the location and potential contacts of MPV17 in the cristae can be obtained. MPV17 appears to be activated by oxidative conditions. Therefore, we will also perform our functional studies under high ROS stress conditions, which may lead to oxidation of the cysteine residues contained in MPV17, resulting in structural changes. In the second part of the study, the three-dimensional structure of MPV17 will be obtained by NMR spectroscopy and cryo-electron microscopy. The monomer present under reducing conditions will be studied by NMR spectroscopy, and the oligomer populated under oxidative conditions will be studied by cryo-electron microscopy in order to detect possible structural changes and thus better understand the mechanism of MPV17 activation. With the obtained structural information, the influence of mutations on MPV17 function can be better understood, which could be the basis for a possible specific therapy of MPV17-induced MDDS.

• DFG Programme: Research Grants