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Daniel Poveda Huertes

Daniel Poveda Huertes

Regulatory mechanisms of mitochondrial biogenesis

Principal investigator: Prof. Dr. Chris Meisinger

Institut für Biochemie und Molekularbiologie
Stefan-Meier-Str. 17
79104 Freiburg

Phone: +49 (0) 761-203-97472
daniel.poveda.huertes@biochemie.uni‐freiburg.de

Abstract

Mitochondria are dynamic organelles and approx. 99% of its proteins are encoded in the nucleus. Therefore they have to be imported as precursor proteins after their translation in the cytosol. Within the organelle they have to fulfill many different functions like the biosynthesis of iron sulphur clusters, heme, lipids, amino acids as well as the respiratory chain and the regulation of apoptosis.

The majority of these precursor proteins are imported via the translocase of the outer membrane (TOM complex) as a main entry gate into the organelle and a translocase in the inner membrane (TIM complex), which mediates transport across the inner membrane.

Around 70% of the precursors have a positively charged targeting signal at the N-terminus, which is recognized by both translocases and targets them finally into the mitochondrial matrix. As a final step of the transport most of this proteins will be processed by the protease MPP that cleaves the presequence thereby generating the mature protein.

The aim of my PhD thesis is to study the role of posttranslational modifications during the import process. This includes irreversible changes like the cleavage of the N-terminal presequence from the precursors proteins but also reversible modifications like phosphorylation. All this modifications have to work together to ensure a dynamic transport of molecules according of the metabolic needs of the cell.

Methods

  • Yeast cell culture
  • Isolation of mitochondria
  • Cell free translation of membrane proteins
  • In vitro import import into mitochondria
  • Blue native PAGE
  • Phos-tag SDS-PAGE and phosphorylation assays
  • Affinity purification