BPS2026 – AI modeling and molecular dynamics reveal a pre-cleavage complex between PINK1 and PARL

Upon import into the mitochondrial inner membrane, the kinase PINK1 is cleaved by the rhomboid protease PARL, followed by retro-translocation and proteasomal degradation. In compromised mitochondria this process fails, leading to PINK1 activation at the outer-membrane and ultimately to mitophagy. Aside from cleavage by PARL, analysis of PINK1 import within intact cells reveals an alternative pathway, directly into the mitochondrial matrix. Both pathways likely proceed via TIM23, in complex with either PARL or Romo1, respectively. AlphaFold modelling predicts that the PINK1 transmembrane domain (TMD) adopts a α-helix when alone or importing via Romo1 for matrix import but shifts to a α/β hybrid when engaged with PARL. In this pre-cleavage complex, PARL appears to partially unfold PINK1 to enable cleavage by the PARL active site residues. Molecular dynamics simulations show how these conformations are stabilized by conserved residues in the import machinery and by PINK1 residues, including several implicated in Parkinson’s disease. We propose that these mutually exclusive complexes determine alternative fates—full-length matrix import versus cleavage and retro-translocation—providing new insight into how PINK1 regulates mitochondrial function and quality control in health and disease.