We use x-ray crystallography, mechanistic biochemistry and protein design techniques to uncover the operating principles of energy-dependent proteolytic machines in mitochondria.
AAA+ molecular machines are used in all organisms to couple the energy of ATP hydrolysis to mechanical work. Unwinding of DNA helices, unfolding of stable proteins, and movement of intracellular cargo all require the generation and specific application of force by AAA+ machines. Given the wide variety of essential cellular processes powered by these enzymes, it is unsurprising that their dysfunction has been implicated in many human diseases, including cancer, diabetes and neurodegeneration.
Our work focuses on the role of AAA+ machines in regulating protein quality control of the mitochondrial inner membrane. Ring-shaped hexameric proteases comprising a AAA+ motor unit linked to a proteolytic chamber, recognize and engage damaged components of the respiratory chain. Cycles of ATP binding and hydrolysis in the AAA+ module drive repetitive conformational motions that enable dislocation of the protein substrate from the membrane, followed by translocation of the unfolded polypeptide into the proteolytic chamber for degradation. Although considerable insight has been gained into the crucial role played by these proteases in maintaining mitochondrial proteostasis, the molecular mechanisms that underlie these functions remain elusive.
Our goal is to understand on a molecular level how mitochondrial energy-dependent proteases select specific protein targets, and the nature of the mechano-chemical coupling that drives dislocation, translocation and ultimately, degradation. Such knowledge will shed light on the general operating principles of the diverse family of AAA+ machines.