Translocation from cytoplasm to mitochondria

• Mitochondria contain sub compartments therefore multiple sorting signals may be required for mitochondrial polypeptides
• Translocation machinery is required for the outer mitochondrial membrane and inner mitochondrial membrane
• Chaperones / hsp70 and signal sequence allows TOM (translocase of outer membrane) to recognise precursor proteins
• MIA is the machinery for import and assembly (disulphide bond relay)
• SAM is the sorting and assembly machinery
• OXA is the insertase / export machinery of the inner membrane
• Translocase of outer membrane (TOM) subunits required for targeting:

  • 40 - central pore
  • 70/20 - peripheral receptors
  • 22 - central receptor
  • 5-7 - assembly / stability
• For cargo recognition by TOM, chaperones (hsp70, hsp90) bind to the precursor protein. This complex binds at TOM 70 / 22 via cochaperones
• Some outer membrane (OMM) polypeptides (beta barrel porins) enter the intermembrane space (IMS) and bind to IMS chaperones (TinyTIMs) which are inserted into OMM by SAM
• Cell free systems can be used to test modifications to signal sequence and requirement for cytosolic or mitochondrial factors e.g. energised mitochondria, cytosolic chaperones, mitochondrial chaperones (using hsp70 mutant mitochondria) - fully folded polypeptides will not be imported
• Translocation of IMS proteins

  1. Polypeptides are transferred from TOM to TIM
  2. Stop transfer signal is released from TIM and is cleaved and released to IMS
  3. Metabolite transporters with internal signals snake through TOM
  4. Metabolite transporters are released to the IMS to bind chaperones
  5. The complex binds to TIM22/54 and then released to the membrane
• Insertion of IMM proteins

  1. Polypeptide is inserted into TIM 23 / 17
  2. N terminal sequence is cleaved
  3. Stop transfer signal directs exit from TIM
  4. Polypeptide enters the matrix via TIM 23 / 17
  5. N terminal sequence is cleaved
  6. Second signal directs the polypeptide to OXA (insertase / export machinery of the inner membrane)
• Signal sequences are cleaved by matrix proteases
• Soluble matrix polypeptides may be passed to hsp60 from mitochondrial hsp70 to fully fold in the matrix
• ATP in mitochondrial import

  1. Import across OMM is driven by ATP hydrolysis by the mitochondria import stimulating factor) and cytosolic and mitochondrial hsp70
  2. TIM further translocates the signal, this is driven by electrochemical hydrogen ion gradient across the IMM and acts on positively charged signaling sequence
  3. The emerging polypeptide is bound by mitochondrial hsp70 with ATP hydrolysis and co chaperones